From: P.Sobieszczuk at Ic.Ac.Uk (Peter Sobieszczuk)

1996 July

From p.sobieszczuk at ic.ac.uk Wed Jul 24 20:01:05 1996 From: p.sobieszczuk at ic.ac.uk (Peter Sobieszczuk) Subject: test only (disregard) Message-ID: test only (disregard)

From csmits at qlt-pdt.com Thu Jul 25 14:01:16 1996 From: csmits at qlt-pdt.com (Claire Smits)

Subject: Stress indicators of mice left in the dark Message-ID:

Hello Transgenic List Subscribers:

Can anyone direct me to any studies, papers, or general information, regarding stress indicators of mice that have been placed in the dark or light reduced areas for 24 hours or longer ?

Thank you

Claire Smits [email protected]

From Allison_F._Treloar at ccmail.bms.com Fri Jul 26 09:23:51 1996 From: Allison_F._Treloar at ccmail.bms.com (Allison F. Treloar)

Subject: Contract knock-out production Message-ID: <[email protected]>

I hope there are enough of you signed up on this new list to help me out..

Aside from Genome Systems, does anyone know of other suppliers of contract knock-out mouse production services. I don't know if DNX has this service up and running yet.

Thanks,

Allison

Technical Supervisor-Transgenics Bristol-Myers Squibb Co. [email protected]

From stewarv at cesmtp.ccf.org Fri Jul 26 11:20:45 1996 From: stewarv at cesmtp.ccf.org (Valerie Stewart M.S.)

Subject: Contract knock-out production -Reply Message-ID:

Allison et. al.--

What do you mean exactly by contract knockout-mouse production? My facility will accept frozen ES cells for injection, to produce chimeras for a fee. We do not do the initial production of the ES cell clones containing the knocked-out gene. We haven't been breeding for germline transmission either, though we may extend the service to that. As of yet, we've only had internal business. Let me know if we can be of assistance.

Valerie Stewart Transgenic/Knockout Facility Cleveland Clinic Research Institute "[email protected]"

From r-carver at nimr.mrc.ac.uk Tue Jul 30 10:11:53 1996 From: r-carver at nimr.mrc.ac.uk (Rick Carver)

Subject: Scientific Meeting on Transgenic Animals Message-ID: <[email protected]>

Dear All

The Laboratory Animal Science Association (LASA) is establishing a specialist Scientific Section on Transgenic Animals. This Section will be launched with an inaugural meeting entitled "Transgenic Animals- Progress, Problems and Potential".

The meeting will be held in London (UK) on Friday 27 September 1996 and has the following programme:

Transgenesis- setting the scene Transgenic animals in Developmental Biology Transgenic animals in Immunology Transgenic animals in Therapeutic Protein Production Transgenic animals in Neuroscience Transgenic animals in Oncology Transgenic animals in Pharmaceuticals Discovery and Development Ethical and legal aspects of the use of transgenic animals

The speakers are from NIMR, ICRF, various University departments and leading biotechnology and pharmaceutical companies.

The cost will be 55 pounds stirling for LASA members and 75 pounds stirling for non-members. Registration forms are available from: LASA Speciality Sections, PO Box 3993, Tamworth, Staffs, B78 3QU. The closing date for registrations is 23 Aug 96.

Rick Carver Head of Biological Services and Institute Veterinarian National Institute for Medical Research The Ridgeway MILL HILL London E-Mail: [email protected] NW7 1AA Tel: + 44 (0)181 959 3666 ext 2199 United Kingdom Fax: + 44 (0)181 913 8601

"There is something fascinating about science: one gets such wholesale returns of conjecture out of such a trifling investment of fact" - Mark Twain

From TSAUNDER at hg-basic1mail.hg.med.umich.edu Mon Jul 29 12:36:47 1996 From: TSAUNDER at hg-basic1mail.hg.med.umich.edu (TSAUNDER)

Alison,

Lexicon Genetics, Inc. makes contract knock-out mice. Their URL is http://www.lexgen.com They are located in Woodlands, Texas (North of Houston). They are using Allen Bradley's ES cell line.

Thom Saunders, Ph.D. Transgenic Animal Model Core Biomedical Research Core Facilities University of Michigan Medical School email: [email protected]

Date: Fri, 26 Jul 1996 08:23:51 -0500 (EST) From: "Allison F. Treloar" Subject: Contract knock-out production To: [email protected] Reply-to: [email protected]

Thanks,

Allison

From p.sobieszczuk at ic.ac.uk Wed Jul 24 19:01:05 1996 From: p.sobieszczuk at ic.ac.uk (Peter Sobieszczuk)

Subject: test only (disregard) Message-ID: test only (disregard)

From csmits at qlt-pdt.com Thu Jul 25 18:01:16 1996 From: csmits at qlt-pdt.com (Claire Smits)

Subject: Stress indicators of mice left in the dark Message-ID:

Hello Transgenic List Subscribers:

Thank you

Claire Smits [email protected]

From Allison_F._Treloar at ccmail.bms.com Fri Jul 26 14:23:51 1996 From: Allison_F._Treloar at ccmail.bms.com (Allison F. Treloar)

Thanks,

Allison

From stewarv at cesmtp.ccf.org Fri Jul 26 15:20:45 1996 From: stewarv at cesmtp.ccf.org (Valerie Stewart M.S.) Subject: Contract knock-out production -Reply

Allison et. al.--

What do you mean exactly by contract knockout-mouse production? My facility will accept frozen ES cells for injection, to produce chimeras for a fee. We do not do the initial production of the ES cell clones containing the knocked-out gene. We haven't been breeding for germline transmission either, though we may extend the service to that. As of yet, we've only had internal business. Let me know if we can be of assistance.

From r-carver at nimr.mrc.ac.uk Tue Jul 30 10:11:53 1996 From: r-carver at nimr.mrc.ac.uk (Rick Carver)

Subject: Scientific Meeting on Transgenic Animals Message-ID: <[email protected]>

Dear All

The Laboratory Animal Science Association (LASA) is establishing a specialist Scientific Section on Transgenic Animals. This Section will be launched with an inaugural meeting entitled "Transgenic Animals- Progress, Problems and Potential".

The meeting will be held in London (UK) on Friday 27 September 1996 and has the following programme:

Transgenesis- setting the scene Transgenic animals in Developmental Biology Transgenic animals in Immunology Transgenic animals in Therapeutic Protein Production Transgenic animals in Neuroscience Transgenic animals in Oncology Transgenic animals in Pharmaceuticals Discovery and Development Ethical and legal aspects of the use of transgenic animals

The speakers are from NIMR, ICRF, various University departments and leading biotechnology and pharmaceutical companies.

The cost will be 55 pounds stirling for LASA members and 75 pounds stirling for non-members. Registration forms are available from: LASA Speciality Sections, PO Box 3993, Tamworth, Staffs, B78 3QU. The closing date for registrations is 23 Aug 96.

From TSAUNDER at hg-basic1mail.hg.med.umich.edu Mon Jul 29 17:36:47 1996 From: TSAUNDER at hg-basic1mail.hg.med.umich.edu (TSAUNDER)

Alison,

Lexicon Genetics, Inc. makes contract knock-out mice. Their URL is http://www.lexgen.com They are located in Woodlands, Texas (North of Houston). They are using Allen Bradley's ES cell line.

Date: Fri, 26 Jul 1996 08:23:51 -0500 (EST) From: "Allison F. Treloar" Subject: Contract knock-out production To: [email protected] Reply-to: [email protected]

Thanks,

Allison

1996 August

From Allison_F._Treloar at ccmail.bms.com Fri Aug 2 16:50:53 1996 From: Allison_F._Treloar at ccmail.bms.com (Allison F. Treloar) Subject: Transgenic Barriers

Hi folks,

I am trying to get a feel for how many institutions are using barrier conditions for their transgenic/knockout colonies. Please reply to me off list and I'll sumarize for the group. Don't forget to include where you're from! no/low barrier = conventional caging and access (+/- filter tops on caging) modified barrier = access restrictions, gowning, isolators, (+/- autoclaving) full barrier = shower in, isolator caging, autoclaved food and bedding

Thanks in advance! Allison Treloar Technical Supervisor - Transgenics Bristol-Myers Squibb

[email protected]

From mgallardo at wsu.edu Fri Aug 2 14:36:49 1996 From: mgallardo at wsu.edu (Mike Gallardo)

Subject: Transgenic Barriers Message-ID: <[email protected]>

At 03:50 PM 8/2/96 -0500, you wrote: >Hi folks, > >I am trying to get a feel for how many institutions are using barrier conditions >for their transgenic/knockout colonies. Please reply to me off list and I'll >sumarize for the group. Don't forget to include where you're from! > >no/low barrier = conventional caging and access (+/- filter tops on caging) > >modified barrier = access restrictions, gowning, isolators, (+/- autoclaving) > >full barrier = shower in, isolator caging, autoclaved food and bedding > >Thanks in advance! >Allison Treloar >Technical Supervisor - Transgenics >Bristol-Myers Squibb > >[email protected] > > >Here at Wegner Hall Vivarium/WSU we use isolator caging, autoclaved food and bedding. We do not shower in, we use steril gowns, booties, mask, hair nets and gloves. > >

(''`-''-/").___..--''"`-._ `o_ o ) `-. ( ).`-.__.`) (_Y_.)' ._ ) `._ `. ``-..-' _..`--'_..- / /--' _. .' (il).-'' (li).' ((!.-' Mike Gallardo BS MS LAT Administative Manager

GO COUGS!!!! Wegner Hall Vivarium G-44 Washington State University Pullman, Wa 99164-6510 Phone (509)335-0977 Fax (509)335-0162 e-mail [email protected] "Brains are for thinking--pencils are for remembering" From fmargoli at umabnet.ab.umd.edu Fri Aug 2 18:13:51 1996 From: fmargoli at umabnet.ab.umd.edu (Frank L. Margolis)

Subject: No subject Message-ID:

Anyone-

Is there an up-to-date master list of knockout and/or transgenics that is maintained anywhere?

Thanks

Frank L. Margolis Ph.D. Department of Anatomy University of Maryland at Baltimore School of Medicine 685 West Baltimore Street Baltimore MD 21201

Phone 410-706-8913 office -8914 lab FAX -2512 Department office

From ignelzi at umich.edu Fri Aug 2 18:15:36 1996 From: ignelzi at umich.edu (Michael A. Ignelzi, Jr.)

Subject: Transgenic Barriers Message-ID:

Modified barrier, access restrictions, gowning, isolators, (+/- autoclaving), Univ of Michigan School of Dentistry (transgenic and knockout colonies)

>Hi folks, > >I am trying to get a feel for how many institutions are using barrier >conditions >for their transgenic/knockout colonies. Please reply to me off list and I'll >sumarize for the group. Don't forget to include where you're from! > >no/low barrier = conventional caging and access (+/- filter tops on caging) > >modified barrier = access restrictions, gowning, isolators, (+/- autoclaving) > >full barrier = shower in, isolator caging, autoclaved food and bedding > >Thanks in advance! >Allison Treloar >Technical Supervisor - Transgenics >Bristol-Myers Squibb > >[email protected] >

From UCIVET at uci.edu Fri Aug 2 15:21:06 1996 From: UCIVET at uci.edu (Clifford R ROBERTS)

Subject: Transgenic Barriers Message-ID: <[email protected]> low barrier: microisolator cages, changed in Class II hood, Masks, lab coats, gloves.

Cliff Roberts Univ Calif, Irvine ______Subject: Transgenic Barriers From: [email protected] at biosmtp Date: 8/2/96 1:19 PM

Hi folks,

[email protected]

From pinkert at cmed.bhs.uab.edu Fri Aug 2 20:17:28 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert)

Subject: Transgenic Barriers Message-ID: <19960803002112601.AAD148@cap1>

> Date: Fri, 02 Aug 96 14:21:06 PST > To: [email protected] > Subject: Re: Transgenic Barriers > Reply-to: [email protected] After pulling up a number of responses to the barrier question - with each individual describing a facility, kindly respond directly to Allison Treloar ([email protected]) as it is getting to be a bit much.

From p.sobieszczuk at ic.ac.uk Sat Aug 3 16:53:41 1996 From: p.sobieszczuk at ic.ac.uk (Peter Sobieszczuk)

Subject: "TgList Backstage" (admin. news *1) Message-ID:

Dear Subscribers,

Welcome to the List, again.

The List is now about 10 days old, and quite frankly, your administrator (listowner) has by now much better understanding for the guys who created e.g. Frankenstain etc., it's easier to create the beast than to control it.

On a positive side: The List has (as of 1st of Aug.) approximately 127 subscribers, surprisingly only, less than 40% from the old rodent- research list.

Congratulation to Claire and Allison for their first and second posting (world debut on day 1 or 2 of the List existence), I will repeat their messages below, since there were only very few people subscribed at that time.

On a technical note: function "reply-to" the list, was set up deliberately to make sending replies easier for people who are not that familiar with the electronics of this game, but as C.A. Pinkert noticed, this could be too much on occasions.

Therefore, an explanation: if you use "reply" function of your e-mail package (e.g. "reply" from "message" menu in Eudora) remember it will go back to the list, not only to the original sender. To send your reply (or any message) just to one person use "new" message or type that address in the "to:" field yourself.

It would also appear to be sensible to cut out everything what is not relevant from the copy of the original letter in your "reply to the list" message.

And try to remember to always fill the subject field in the new postings as well as signing your message (makes life easier for everyone).

Hope this will be of some help, and wishing you happy postings.

Peter [email protected]

Copies of the first two posting from the last week: ______Hello Transgenic List Subscribers:

Thank you

Claire Smits [email protected] ______I hope there are enough of you signed up on this new list to help me out..

Thanks,

Allison

Technical Supervisor-Transgenics Bristol-Myers Squibb Co. [email protected] ______From MEISLERM at hg-basic1mail.hg.med.umich.edu Sat Aug 3 14:04:43 1996 From: MEISLERM at hg-basic1mail.hg.med.umich.edu (MEISLERM)

Subject: brain and neuron specific promoters Message-ID:

Does anyone know of a recent review of promoters with in vivo specificity for subsets of neurons, glia, or brain regions? We are looking for promoters specific for brain regions, spinal cord, motor neurons, etc. Thanks for your suggestions.

Miriam Meisler, Ph. D. Department of Human Genetics University of Michigan 4708 Medical Sciences II Ann Arbor, MI 48109-0618 tel 313 763 5546; FAX 313 763 9691.email: [email protected]

From gstuart at UVic.CA Sat Aug 3 18:01:42 1996 From: gstuart at UVic.CA (Greg Stuart)

Subject: Rat liver anatomy question Message-ID: <[email protected]>

Hello - I'd like to draw on the collective experience of the subscribers. Can anybody describe where the following regions of the rat liver are located?

- centrilobular region - midzonal hepatocytes - periportal regions

Also, are these regions found in each lobe of the liver, or are they specific to particular lobes or regions? Please respond directly, thanks. Cheers, Greg :)

-- Greg Stuart [email protected] Centre for Environmental Health . .*#*. .*#*. .*#*. Department of Biology * | | | * | | | * * | | | * | | | * University of Victoria * | | | * * | | | * * | | | * * | | | * P.O. Box 3020 * | | | * * | | | * | | | * * | | |* Victoria, B.C., CANADA `*#*' `*#*' `*#*' `*#*' V8W 3N5. Tel (604)472-4067, Fax(604)472-4075 http://darwin.ceh.uvic.ca/students/gstuart/

From hage at med.unc.edu Sat Aug 3 23:01:14 1996 From: hage at med.unc.edu (John Hagaman)

>Hi folks, > >I am trying to get a feel for how many institutions are using barrier >conditions >for their transgenic/knockout colonies. Please reply to me off list and I'll >sumarize for the group. Don't forget to include where you're from! we us modified barrier = access restrictions, gowning, isolators, (+/- autoclaving, both )

> >[email protected] >

John R. Hagaman Pathology Department University of North Carolina Medical School 7525 Brinkhous-Bullitt, Room 703 Chapel Hill, N.C. 27705

966-6912 office 966-8800 Fax

From kelly at citi2.fr Mon Aug 5 15:12:58 1996 From: kelly at citi2.fr (U344)

Subject: mouse testosterone levels Message-ID:

Hello, does anyone know of a way to measure mouse testosterone levels, or of anyone who is? I'd really like to avoid having to set up an ELISA for a few samples... cheers...

INSERM Unite 344 Endocrinologie Moleculaire 156 rue de Vaugirard, 75730 Paris cedex 15 France

'Man... I don't EVEN have an opinion...'

From browng at medicine.wustl.edu Mon Aug 5 08:53:29 1996 From: browng at medicine.wustl.edu (Gary Brown)

Subject: WWW resource Message-ID:

Just a brief wee message here from St Louis. At the risk of tooting my own horn I'd like to let anyone interested know about my microinjection / transgenisis related homepage. Apologies go to those who may have seen this on the COMPMED listings, or to those who subscribe to this list after my listing THIS list on the page in question! Anyways, here it is...

Page Name: The Microinjection Workshop Page address: http://ourworld.compuserve.com/homepages/TheBroons/homepage.htm How best to find: Although I have registered with most of the widely used search engines, Yahoo is still a problem.. Best to use the Starting Point (www.stpt.com) and select the combination Metacrawler engine searching on "microinjection".

I'm trying to build up an E-mail mailing list to alert when the page is updated (approx. 1 x per month). If interested please reply OFF-LIST tothe following:

[email protected]

Thanx :-)

Gary From Allison_F._Treloar at ccmail.bms.com Mon Aug 5 10:59:37 1996 From: Allison_F._Treloar at ccmail.bms.com (Allison F. Treloar)

Subject: Summary: Tg Barriers Message-ID: <[email protected]>

First and foremost, a big thanks to everyone who responded to my question regarding barrier conditions for trangenic colonies. Here's the outcome:

17 respondents 3 facilties use low/no barrier conditions 10 facilities use modified barrier conditions 4 facilities use full barrier conditions low/no barrier respondents stated that these conditions were used primarily for single transgenic lines or areas where investigators could generally access transgenic stock animals. modified barrier users seem to rely on access restrictions, gowing, dedicated staff, isolator caging and autoclaved food/bedding, but don't go as far as shower-in or sterile clothing/scrubs. Most cage changing/animal manipulation is done in laminar flow hoods. This is certainly the most popular, flexible and cost effective option based on current opinion. full barriers were used by a few facilities and included air showers, sterile clothing/scub requirements as well as all the modified barrer conditions. No respondents with full barriers used shower-in.

Thanks, again for all your help.

Allison

[email protected]

From gina_seeburger at Merck.Com Mon Aug 5 15:07:08 1996 From: gina_seeburger at Merck.Com (Gina Seeburger)

Subject: "TgList Backstage" (admi Message-ID: <199608052001.QAA29238@igw2>

RE>"TgList Backstage" (admin.

------Date: 8/3/96 10:56 AM To: Gina Seeburger From: [email protected] Dear Subscribers,

Peter [email protected] P.S.

Thank you

Thanks,

Allison

Technical Supervisor-Transgenics Bristol-Myers Squibb Co. [email protected] ______

From TSAUNDER at hg-basic1mail.hg.med.umich.edu Tue Aug 6 10:57:55 1996 From: TSAUNDER at hg-basic1mail.hg.med.umich.edu (TSAUNDER)

Subject: Meeting Announcement Message-ID:

THE UNIVERSITY OF MICHIGAN CENTER FOR ORGANOGENESIS 1st International Symposium

Molecular Control of Organogenesis

Saturday, October 5, 1996 Horace H. Rackham Graduate School Amphitheater The University of Michigan Ann Arbor, Michigan

8:45-9:00 Opening Remarks, Dr. Giles G. Bole, Dean, The University of Michigan Medical School Dean

9:00-9:50 Dr. Eric Olson, University of Texas, Dallas, Texas USA, "Genetic control of myogenesis"

9:50-10:40 Dr. Peter Gruss, Max Planck Institute of Biophysical Chemistry, Germany, "Genes required for the development of the visual system"

10:40-10:50 Break

10:50-11:40 Dr. Cliff Tabin, Harvard Medical School, Boston, Massachusetts, USA, "Signals patterning the vertebrate limb"

11:40-12:30 Dr. Yuh Nung Jan, University of California, San Francisco, California, USA, "Molecular control of neural development"

12:30-2:30 Poster Session (lunch provided)

2:30-3:20 Dr. Paul Sternberg, California Institute of Technology, Pasadena, California, USA, "Precise formation of a hole: the C. elegans vulva"

3:20-4:10 Dr. Richard M. Harland, University of California, Berkeley, California, USA, "Re-use of signaling molecules in vertebrate axis formation and organogenesis"

4:10-4:20 Closing Remarks, Dr. Seigo Izumo, Acting Director, Center for Organogenesis, The University of Michigan

Friday, October 4, 1996 5:30-7:00 p.m.Meet the Speakers at an Open Reception, Museum of Art, The University of Michigan

For further information, please contact Michelle Shukait at 936-2499 or [email protected] From hage at med.unc.edu Wed Aug 7 00:28:28 1996 From: hage at med.unc.edu (John Hagaman)

Subject: White noise for Mouse Embryo Yield Message-ID:

Does anyone have experience re: noise in mouse rooms and how it may affect the yield of embryos? Do mice "get used to certain noises" Does with drawal or introduction of noise change the yield?

Responses off list will be summarized.

Thanks

966-6912 office 966-8800 Fax

From fmargoli at umabnet.ab.umd.edu Wed Aug 7 10:38:50 1996 From: fmargoli at umabnet.ab.umd.edu (Frank L. Margolis)

Subject: Knock out master list Message-ID:

Thanks to all for your responses to my query, they were most helpful.

Frank Margolis

Phone 410-706-8913 office -8914 lab FAX -2512 Department office From r-carver at nimr.mrc.ac.uk Thu Aug 8 17:47:28 1996 From: r-carver at nimr.mrc.ac.uk (Rick Carver)

Subject: LASA Transgenics Meeting Message-ID: <[email protected]>

Dear All

I am now able to give details of the speakers for the forthcoming LASA Transgenics meeting.

Transgenesis- setting the scene Dr Roy Forster, Transgene Transgenic Animals in Developmental Biology Dr Reter Rigby, NIMR Transgenic Animals in Immunology Dr Mike Owen, ICRF Transgenic Animals in Production Proteins Dr Ron James, Pharmaceutical Proteins Ltd Transgenic Animals in Neuroscience Prof Rick Lathe, Univ of Edinburgh Transgenic Animals in Oncology Dr Frances Balkwell, ICRF Transgenic Animals in Pharmaceuticals Dr John McNeish, Pfizer US Discovery and Development Ethical and Legal Aspects of the Use of Dr Kenneth Boyd, Univ of Edinburgh Transgenic Animals

The cost will be 55 pounds stirling for LASA members and 75 pounds stirling for non-members. Registration forms are available from: LASA Speciality Sections, PO Box 3993, Tamworth, Staffs, B78 3QU, UK. The closing date for registrations is 23 Aug 96.

From gac at po.mri.montana.edu Wed Aug 7 18:36:49 1996 From: gac at po.mri.montana.edu (George Carlson)

Subject: discontinuation of strains Message-ID:

Most of the intra-H-2 recombinant B10 congenic strains developed by the late Dr. Jack Stimpfling of McLaughlin Research Institute will be discontinued as live stock in the near future. Following Dr. Stimpfling's retirement, a grant from the American Cancer Society allowed us to complete isolation of his more recently identified recombinant chromosomes, to refine localization of the crossover sites within H-2, to delimit the congenic intervals for each strain, and to preserve the strains for future use. We are now in the final stage of this project and expect to have all strains frozen before January 1997. DNA samples will continue to be available.

The strains are listed in: Turner et al. Meiotic recombination within the H-2K--H-2D interval: characterization of a panel of congenic mice, including 12 new strains, using DNA markers. Immunogenetics 1993, 38:332-340.

Additional data on our congenic panel are being prepared for publication.

Breeding pairs are available for any investigator who requests them. We only ask that investigators who "adopt" a strain make it available to others in the community and provide notification if they stop the line.

I will provide a listing of the strains that we will continue as live stock after we get a better idea of interest. Please let me know if you need any of these mice. Suggestions of other appropriate lists for posting this notice would also be appreciated.

Thank you.

George A. Carlson ([email protected]) McLaughlin Research Institute 1520 23rd Street South Great Falls, Montana 59405 406 452 6208, fax: 454 6019 or 454 6004

From S.Tan at Anatomy.Unimelb.EDU.AU Thu Aug 8 20:41:53 1996 From: S.Tan at Anatomy.Unimelb.EDU.AU (Seong Seng Tan)

>Does anyone have experience re: noise in mouse rooms and how it may affect >the yield of embryos? Do mice "get used to certain noises" Does with >drawal or introduction of noise change the yield? >

We are currently undergoing major building renovations and this week alone, three of our litters have been eaten by their mothers. We suspect loud sudden noises are responsible.

Seong-Seng Tan

======Senior Lecturer, Department of Anatomy & Cell Biology The University of Melbourne Parkville 3052 Victoria, Australia Fax 61 3 9 347 5219, Phone 61 3 9 344 5787 Web: http://www.anatomy.unimelb.edu.au

======

From fmargoli at umabnet.ab.umd.edu Thu Aug 8 11:11:34 1996 From: fmargoli at umabnet.ab.umd.edu (Frank L. Margolis)

Subject: Lists Message-ID:

To all-

In response to several requests I am posting a summary of all the info I have received about the availability of lists of Knockouts and transgenics and specific strains (without any editorial evaluation).

BioMedNet has at least a partial list of knockouts

JAX homepage is http://www.jax.org

Current list of induced mutant strains accepted for inclusion in the Induced Mutant Resource (IMR) at JAX is on the IMR home page- http://lena.jax.org/resources/documents/imr/

An index of strains is available at- http://lena.jax.org/resources/documents/imr/imr.html and more details at- http://lena.jax.org/resources/documents/imr/imrdata.html for queries about any JAX mice contact- [email protected] Also the following web sites have info that may prove to be useful- NIH transgenic directory Tbase http://www.gdb.org/Dan/tbase/tbase.html

Mouse and rat research home page- http://www.cco.caltech.edu/~mercer/htmls/rodent_page.html#homepages and as was also pointed out to me: 1-a Medline search for info on specific genes or strains as well as 2- right here where I started all this.

Thanks again for all your input.

Frank Margolis

From gstuart at UVic.CA Thu Aug 8 08:37:43 1996 From: gstuart at UVic.CA (Greg Stuart)

Subject: Isolation of bone marrow & colon tissues Message-ID: <[email protected]>

Hello everyone. Does anybody have protocols that they could share for the isolation of rat (or rodent) bone marrow and colon tissue? Thanks, Greg :)

-- Greg Stuart [email protected] Centre for Environmental Health . .*#*. .*#*. .*#*. Department of Biology * | | | * | | | * * | | | * | | | * University of Victoria * | | | * * | | | * * | | | * * | | | * P.O. Box 3020 * | | | * * | | | * | | | * * | | |* Victoria, B.C., CANADA `*#*' `*#*' `*#*' `*#*' V8W 3N5. Tel (604)472-4067, Fax(604)472-4075 http://darwin.ceh.uvic.ca/students/gstuart/ From r-lovell at nimr.mrc.ac.uk Fri Aug 9 18:38:45 1996 From: r-lovell at nimr.mrc.ac.uk (Robin Lovell-Badge)

Subject: Copy of: Re: White noise for Mouse Embryo Yield Message-ID:

Dear All

There is considerable published and anecdotal stuff on noise in animal rooms. Excessive noise, e.g. from building works, can have all sorts of effects which will lower reproductive efficiency. Mice seem to particularly dislike sudden noises, so it can be beneficial to have music playing (classical or modern, but not too much rap !) during the day when cages are being changed, etc. But again, it should not be very loud, just enough to mask the dropping of cages, screams of students, etc.

If anyone is desperate I can try to locate published work on this - but it is not recent and may take a while.

Robin Lovell-badge

>You wrote: > >>Does anyone have experience re: noise in mouse rooms and how it may affect >>the yield of embryos? Do mice "get used to certain noises" Does with >>drawal or introduction of noise change the yield? >> > > While working at the University of Kentucky, I found a number of >interuptions in breeding colonies due to the noise caused by construction. >Liters were either canabalized by the mother or breeding stopped >completely. Hope this helps. > > >------> ( )_( ) > Richard S. Cluck, B.S., LATG o o > Supervisor, VMU, VA, Lexington ==\o/== > Research 151 /\ ) > Cooper Drive / \ ( > Lexington, KY 40511 / \) > Phone: 606-281-4927 > Fax: 606-281-4989 > E-Mail: cluck,[email protected] >------From cfoltz at MIT.EDU Mon Aug 12 12:50:40 1996 From: cfoltz at MIT.EDU (Charmaine Foltz )

Subject: control of P. pneumotropica Message-ID: <[email protected]>

I am currently treating a small colony of Rag-2 mice to try to eliminate Pasteurella pneumotropica, which has been a significant pathogen for this particular genotype. My question to the group is how many of you have had experience with doing this (I am using enrofloxicin in the water). Additionally, having read recently that humans can colonize with the bacteria (Cudrado-Gomez LM, et al. Pasteurella pneumotropica pneumonia in a patient with AIDS. Clinical Inf. Dis, 1995;21:445-6), I wonder if anyone on the list has had experience with a barrier facility becoming colonized with the agent, presumably from contamination by a human source.

Thank you, Charmaine Foltz, DVM, Dipl. ACLAM MIT Div. Comparative Medicine 77 Massachusetts Ave. Bldg. 45 Cambridge, MA 02139 (617)252-1804 (617)258-5708 (fax) [email protected]

From dknudsen at scruznet.com Mon Aug 12 21:32:45 1996 From: dknudsen at scruznet.com (Dave Knudsen)

Subject: control of P. pneumotropica Message-ID:

>I am currently treating a small colony of Rag-2 mice to try to eliminate >Pasteurella pneumotropica, which has been a significant pathogen for this >particular genotype. My question to the group is how many of you have had >experience with doing this (I am using enrofloxicin in the water). >Additionally, having read recently that humans can colonize with the >bacteria (Cudrado-Gomez LM, et al. Pasteurella pneumotropica pneumonia in >a patient with AIDS. Clinical Inf. Dis, 1995;21:445-6), I wonder if anyone >on the list has had experience with a barrier facility becoming colonized >with the agent, presumably from contamination by a human source.

Charmaine -

I've had some experience with this agent. In a colony of over 12,000 SCID mice used in transplantation studies (and therefore routinely irradiated), we were highly motivated to remove pasteurellosis from the list. Using draconian barrier procedures and caesarian rederivation of every strain to be introduced into the facility, we were able to rid all of our colonies of P pneumo as well as everything else I know how to test for. Individually ventillated caging, a frequent health monitoring system, and restricted access help kept it that way. I'll be happy to give you more details off line should you be interested in this approach.

Dave Knudsen DVM, MS, DACLAM Scotts Valley, California, USA

*** "In my judgement it is either an enigma or some kind of bug. If it dies, I will take it apart and see what its arrangements are. I never had a thing perplex me so." Mark Twain, 1893 (from "Extracts from Adam's Diary") ***

From dbowtell at petermac.unimelb.edu.au Tue Aug 13 21:49:47 1996 From: dbowtell at petermac.unimelb.edu.au (David Bowtell)

Subject: Double KO with increased G418 Message-ID:

We have used increased G418 concentration as a means of selecting homozgous cells on two occassions. One construct had a pGKNeo casssette and the cells were still laughing in 4mg/ml (we blinked first). The second construct was promoterless, the cells were marginally G418 resistant and it worked like a dream. The paper by Mortensen (MCB 12, 2391) used a pGKNeo cassette but I am unclear whether it was the point mutant (low activity) neo cassette. So we would be interested in other people's experience with this approach to double KO in ES cells (and any tidbits on experience targeting the second allele indepndantly).

Many thanks.

David Bowtell.

******* Please note new email address ********** David Bowtell Principal Research Fellow Research Division Peter MacCallum Cancer Institute Locked Bag 1 A'Beckett St, Melbourne 3000

Lab 61-3-96561287 Office 61-3-96561296 Fax 61-3-96561411 email: [email protected]

From wixson at biovax.dnet.basf-ag.de Tue Aug 13 20:16:55 1996 From: wixson at biovax.dnet.basf-ag.de (Sally Wixson, VMD, MS)

Subject: Knockout advice Message-ID: <[email protected]>

I direct the Lab Animal department of a pharmaceutical research company that has an active, but disorganized, knock out team. I have been here eight months and trying to get their animal work back on track. I would be grateful for your advice on several topics: .....what strain of vactomized males do you use and why? How do you assess their sterility (other than ordering then vasec. by the vendor)? How often do you use vasec. males? (my invesitagors use each male once/week or less!)

.....what strain of embryo recipients do you use for ES cell work? Our donors are C57BL/6, recipients are B6D2F1's. My team wants a "big" target so they keep the females here three months or so on a high fat diet so they are big enough to inject. Obviously, this leads to a waste of cage space. Many people use ICR's or CD1's as recipients because they are a big target and good moms!

.....in breeding chimeras, our PI insists that each chimera must produce100 pups (black) before being considered a failure at germ line transmission. Many of the chimeras are lowly sterile or sterile, so this format entails breeding formany months to more than a year. Several other labs told me their chimeras get three matings and if no agouti pups are produced, they are ENA'd.

.....do you produce your own ES cells or get them from donation/purchase from other labs? We are currently buying the RW4 ES cell line from Genome Systems. Do you require MAP testing of the new ES cell lines?

.....do you superovulate your females for embryo donation? My PI says this doe not yield viable embryos for ES cell work, so they breed 4-5X the number of females they rpedict will be needed for each injectionso that they have enough embryos.

Many thanks for the advice, Sally K, Wixson, VMD, MS

From wixson at biovax.dnet.basf-ag.de Tue Aug 13 20:16:55 1996 From: wixson at biovax.dnet.basf-ag.de (Sally Wixson, VMD, MS)

.....what strain of embryo recipients do you use for ES cell work? Our donors are C57BL/6, recipients are B6D2F1's. My team wants a "big" target so they keep the females here three months or so on a high fat diet so they are big enough to inject. Obviously, this leads to a waste of cage space. Many people use ICR's or CD1's as recipients because they are a big target and good moms! .....in breeding chimeras, our PI insists that each chimera must produce100 pups (black) before being considered a failure at germ line transmission. Many of the chimeras are lowly sterile or sterile, so this format entails breeding formany months to more than a year. Several other labs told me their chimeras get three matings and if no agouti pups are produced, they are ENA'd.

Many thanks.

David Bowtell. ******* Please note new email address ********** David Bowtell Principal Research Fellow Research Division Peter MacCallum Cancer Institute Locked Bag 1 A'Beckett St, Melbourne 3000

From dknudsen at scruznet.com Mon Aug 12 21:32:45 1996 From: dknudsen at scruznet.com (Dave Knudsen)

Charmaine -

From cfoltz at MIT.EDU Mon Aug 12 12:50:40 1996 From: cfoltz at MIT.EDU (Charmaine Foltz )

From browng at medicine.wustl.edu Wed Aug 14 08:52:59 1996 From: browng at medicine.wustl.edu (Gary Brown)

Subject: Knockout advice In-Reply-To: <[email protected]> Message-ID:

Hi, Just throwing in my 2 cents worth...

On Tue, 13 Aug 1996, Sally Wixson, VMD, MS wrote:

> .....what strain of vactomized males do you use and why? How do you assess > their sterility (other than ordering then vasec. by the vendor)? How often do > you use vasec. males? (my invesitagors use each male once/week or less!)

I use Swiss Webster VAS-X males, ordered in from Taconic Farms. In my previous position we did our own vasectomies on C57Bl/6J males (pronuclear microinjection only) and assessed their sterility by co-housing with Swiss Webster (or other inexpensive white mouse strain) females (2) for approx. 6 weeks prior to use. Males were vasectomized at 4-5 weeks of age, excising a minimum of 1cm of vas deferens per side. > > .....what strain of embryo recipients do you use for ES cell work? Our donors > are C57BL/6, recipients are B6D2F1's. My team wants a "big" target so they keep > the females here three months or so on a high fat diet so they are big enough > to inject. Obviously, this leads to a waste of cage space. Many people use > ICR's or CD1's as recipients because they are a big target and good moms!

I have used B6D2F1s for recipients although now use Swiss Webster (occasionally ICR) females as they are a relatively large mouse strain. As for feeding them on a high fat diet I personally do not subscribe to the 'big target' philosophy - at least no bigger than using regular mouse chow for my chosen strain. Since we have a pool of SW females from which to pick those in estrus and individuals may be passed over several times, high fat diet pushes some of them to a size where surgery becomes more difficult due to the increased fat deposits around the oviduct/ovary/UT junction, and I believe that the increased incision size in the body wall that this necessitates increases the risk to the animal. > > .....in breeding chimeras, our PI insists that each chimera must produce100 > pups (black) before being considered a failure at germ line transmission. Many > of the chimeras are lowly sterile or sterile, so this format entails breeding > formany months to more than a year. Several other labs told me their chimeras > get three matings and if no agouti pups are produced, they are ENA'd. This is insane! My initial reaction would be to tell your PI to get a life! Our policy is to discard all female chimeras (low germline transmission, increased sterility in the F1 generation when successful) and to discard all >50% (by color coat) male chimeras. 3 matings (strikes) and you're out sounds appropriate to me. > > .....do you produce your own ES cells or get them from donation/purchase from > other labs? We are currently buying the RW4 ES cell line from Genome Systems. > Do you require MAP testing of the new ES cell lines?

Our lab has approached other labs for ES cells, although we are trying to establish our own cell line currently. With reference to the RW4 cell line, I have used this effectively on several occasions and note that it has a high color coat conversion rate. I also know the individual who created this cell line, and can forward any questions you may have to her. Our institution requires MAP testing of all ES cell lines for use in mice. > > .....do you superovulate your females for embryo donation? My PI says this doe > not yield viable embryos for ES cell work, so they breed 4-5X the number of > females they rpedict will be needed for each injectionso that they have enough > embryos. > Yes, I superovulate females for embryo donation. I typically superovulate 7 C57Bl/6J females for one day's effort, with a variation between 20-25 morula per plugged female. I culture these in the incubator overnight to blastocysts with an 85-95% conversion rate. The females used are between 3.5-4.5 weeks old. I use Sigma Chemical Co. new embryo tested PMSG and HCG. My stud males are mated no more than once weekly. There is a reference for superovulating females for blastocyst production, but I would have to look it up on Medline. Sounds to me like your PI needs to do some homework!

Hope this helps!

Gary Brown E-mail: [email protected] or [email protected] Web: http://ourworld.compuserve.com/homepages/TheBroons/ "The Microinjection Workshop"

From kveen at nki.nl Thu Aug 15 12:52:09 1996 From: kveen at nki.nl (Karin van Veen) Subject: blastocyst culture Message-ID:

Hi, you all!

I hope you can help me. At our institute we do zygote injections and blastocyst injections. For some time now we have some problems with the blastocyst yields. It is either a superovulation problem or a flushing problem (or both). Can someone give me some tips concerning the time schedule on the superovulation. In what time intervals do you inject the hormones, and isolate the embryo's? We tried out a lot, but can not improve our yield. We use C57/bl6 mice as donors. We are also thinking, when it should be a flushing problem, to isolate the zygote on day 0,5, and culture them into blastocyst to be injected. Has anyone experience on this? How long do you have to culture the zygotes to get blastocysts, are they good to be injected with ES cells? All tips are welcome!

Karin. The Netherlands Cancer Institute Amsterdam

From browng at medicine.wustl.edu Thu Aug 15 08:23:30 1996 From: browng at medicine.wustl.edu (Gary Brown)

Subject: blastocyst culture In-Reply-To: Message-ID:

On Thu, 15 Aug 1996, Karin van Veen wrote: > Can someone give me some tips concerning the time schedule on the > superovulation. In what time intervals do you inject the hormones, and > isolate the embryo's? > We use C57/bl6 mice as donors.

We use a 12hr/12hr light-dark cycle from 4am to 4pm in our facility, coupled with 47hrs between PMSG and HCG. These hormones are typically given at 10.30am and (47hrs later) 9.30am repectively for this light cycle. We use hormones obtained through Sigma Chemical Co. (Embryo culture tested - new product) and have been obtaining yields of MORULAE at a rate of 20-25 per plugged C57Bl/6J female. These morulae are cultured overnight to blastocysts for ES cell injection in Gibco's modified BMOC-3 culture media with pen/strep.added in a 37deg C 5% CO2 incubator.

> We are also thinking, when it should be a flushing problem, to isolate the > zygote on day 0,5, and culture them into blastocyst to be injected. Has > anyone experience on this? How long do you have to culture the zygotes to get > blastocysts, are they good to be injected with ES cells? > All tips are welcome! > For culture past day 0.5, you would need 3 more days and some consistently good media. I suggest you flush morulae! Although I doubt that flushing would present too much of a problem, this is how I do it. Using a short bevel stainless 30g hypo., introduce the needle at the last oviduct loop before the UT junction and flush media through this area expelling the embryos out of the uterus (cut 1cm down from the oviduct). If this is problematical, flush through the infundibulum (requires a little more finesse).

Hope this helps out!

Gary Brown Email: [email protected] or [email protected] Web: http://ourworld.compuserve.com/homepages/TheBroons/ "The Microinjection Workshop"

From m.hampson at ic.ac.uk Thu Aug 15 18:18:40 1996 From: m.hampson at ic.ac.uk ([email protected])

Subject: Test - please ignore Message-ID:

Test - please ignore

From gina_seeburger at Merck.Com Thu Aug 15 16:15:46 1996 From: gina_seeburger at Merck.Com (Gina Seeburger)

Subject: blastocyst culture Message-ID: <[email protected]>

REPLY o REPLY o REPLY o REPLY RE>blastocyst culture Karin, We were having problems too here in the US, but giving the first injection(PMSG, Sigma brand) between 3:00pm and 4 (.3cc, .7.5IU) and the second (hCG) around 1:00 seems to help. We are also using C57BL/6 donors for blast injections. Collected around 7:30am. Also summer seems to create a low. Good luck Gina Seeburger Merck & Co,Inc. [email protected]

------Date: 8/15/96 5:57 AM To: Gina Seeburger From: [email protected]

Hi, you all!

From tjf at uci.edu Fri Aug 16 13:11:55 1996 From: tjf at uci.edu (Tom Fielder)

Subject: oocyte injection Message-ID:

I am brand new to the field of transgenics, and I am setting up a transgenic core facility at UC-Irvine. I am still trying to optimize my pipets for injection of DNA into oocytes. I'm using a Sutter P-97 puller, and 1mm OD x 0.78mm ID capillaries with internal filament. I got some pipets that seemed to work very well yesterday, judging by the swelling of the pronucleus and no visible cell contents leaking out after withdrawing the pipet, but this morning 65% of the injected cells had died (cytoplasm was condensed and dark brown), while none of the uninjected controls looked like this. I'm afraid that the taper of the needle is too abrupt near the tip (i.e., it gets too big too fast). A very patient person at Sutter is helping me with the pipets, but my question for the list is this: Is the condensed, dark brown cytoplasm indicative of mechanical trauma, or is it a result of bad DNA/buffer, or something else? A second question: The books recommend using females that are 24-25 days old for superovulation. What happens if you use older ones? Will the yield of oocytes be significantly less? I've been using 6-8 week old FVB mice and getting about 30 eggs/mouse, but I've noticed that a large number of them aren't fertilized, even from the females that were obviously plugged. Any thoughts? Tom

Thomas J. Fielder UC-Irvine University Lab Animal Resources Transgenic Mouse Facility 147 BSA Irvine, CA 92697-1310 e-mail: [email protected] phone: 714-824-8579 FAX: 714-824-2003

From MKENNEDY at chmeds.ac.nz Sat Aug 17 23:12:42 1996 From: MKENNEDY at chmeds.ac.nz ([email protected])

Subject: Double KO with increased G418 Message-ID: <[email protected]>

Date sent: 17-AUG-1996 22:06:32

Hi Dave,

Just got back from this year's Queenstown Meeting, which (for my sins) I organised.

>We have used increased G418 concentration as a means of selecting homozgous >cells on two occassions. One construct had a pGKNeo casssette and the cells >were still laughing in 4mg/ml (we blinked first). The second construct was >promoterless, the cells were marginally G418 resistant and it worked like a >dream. The paper by Mortensen (MCB 12, 2391) used a pGKNeo cassette but I >am unclear whether it was the point mutant (low activity) neo cassette. So >we would be interested in other people's experience with this approach to >double KO in ES cells (and any tidbits on experience targeting the second >allele indepndantly).

For what it is worth, I sequenced Mortensen's pNTK construct, which I am pretty sure was the one he used in the MCB paper, and it was of the mutant neo variety. I am at home at present, but if you haven't seen it, then John Sedivy had a very interesting MCB a year or so back on titrating G418 to obtain high efficiency KO's, with some interesting observations that related to the homozygous KO method. Let me know if you need more info, and I'll dig it out when I am at work. I have an oligo from close to the polymorphic site in the neo gene, if you want to check other neo constructs by sequencing - you're welcome to some of it.

Cheers,

Martin

NNNN NN Martin A Kennedy (E-mail = [email protected]) ZZZZZZZ NN NN NN Cytogenetic and Molecular Oncology Unit ZZZ NN NN NN Christchurch School of Medicine ZZZ NN NNNN Christchurch, New Zealand ZZZZZZZ Phone (64-3)364-0880 Fax (64-3)364-0750

From stewarv at cesmtp.ccf.org Mon Aug 19 12:40:44 1996 From: stewarv at cesmtp.ccf.org (Valerie Stewart M.S.)

Subject: Knockout advice -Reply Message-ID:

Hi Sally--

We use Swiss Webster vasectomized males, bought from Taconic. We don't routinely test them, and have so far (knock on wood!) never had a leak. When I used to do the vasectomies myself, we tested them with two females each, let them plug both, then waited 3 weeks for hopefully no resultant pregnancies. We use vasectomized males twice a week to every other day. They can be used every night for a couple of weeks before needing some down time. Once a week is ridiculous, they don't need to keep their sperm count up after all! We use them for a year as well, or until they stop performing. We use Swiss Webster or CD-1 females for recipients--they are easy to estrus select, good mothers, and cheap. I wouldn't keep them on a high fat diet--in fact, we routinely cull out recipients that are too fat and discard them.

As for testing germline transmission, 100 pups to "prove" lack of transmission is the standard, but practicality usually dicatates compromise. We only breed males over 50% chimeric, females only if there are no male chimeras available. Three litters is a good number, if they're good-sized litters, I usually try for 25-30 pups.

We are using E14.1 cells from Cincinnati, but I've used CCE, D3, J1, R1, etc. Cell line is crucial; I've heard Genome Systems' line works fine.

We superovulate for blastocysts routinely, using C57Bl6 females. We generally do 10 at a time, with the standard amounts/timing of hormones, and a 6:30am/6:30pm light cycle. The one thing we find is crucial is to match body weight of the donors, 13-15g at time of PMS administration. We do three injection days per clone, averaging 30 blastocysts per 10 donors (with the occasional null day). I have gotten good results with number and % chimerism, as well as germline transmission with these blastocysts. We only use the males once per week, and usually only for 6-8 months, though a colleague says she uses them twice a week for a year with no trouble.

Good Luck-- Valerie Stewart Transgenic/Knockout Facility Cleveland Clinic Research Institute "[email protected]"

>>> Sally Wixson, VMD, MS - 8/13/96 1:16 PM >>> I direct the Lab Animal department of a pharmaceutical research company that has an active, but disorganized, knock out team. I have been here eight months and trying to get their animal work back on track. I would be grateful for your advice on several topics: .....what strain of vactomized males do you use and why? How do you assess their sterility (other than ordering then vasec. by the vendor)? How often do you use vasec. males? (my invesitagors use each male once/week or less!)

From crussell at ResGen.COM Mon Aug 19 12:03:24 1996 From: crussell at ResGen.COM (Chris Russell)

Subject: employment opportunity Message-ID: <[email protected]>

Position Available Immediately! Lab Animal Technologist/Research Assistant

Skills Required: --Knowledgeable in theoretical embryology --Skilled in applied embryology and microsurgery techniques --Primarily laboratory work using the mouse as a model system --Molecular biology experience preferred

--Salary: $28-30K plus great benefits

--Excellent work environment in a growing biotech firm in Huntsville, AL

Please send resume and references to: Dr. Chris Russell Research Genetics, Inc. 2700 Memorial Parkway, SW Huntsville, AL 35801 1-800-711-2089 e-mail: [email protected] *********************************************************************** **** Christopher G. Russell, Ph.D. RESEARCH GENETICS, INC Resources for Research -_+ _+_+ ++-_ _+-_ _+_ _++ _+-_ -+ -+ -+ +- +- -+ + -+ -+ -+ + -+ -+ -+ -+ +- -+ -+ -+ -+ -+ -+ -+ -+ -++-+ -+_-+ -+_+ -++- -+_++ -+_+ 2700 Memorial Parkway SW Huntsville, AL 35801 Phone 1-800-533-4363, ext 2211 or Direct 1-800-711-2089 U.K. 0-800-89-1393 Fax 1-205-551-1021 or 1-205-536-9016 [email protected] *********************************************************************** ******

Subject: blastocyst culture -Reply Message-ID:

Karin--

We use 5 units PMS and HCG administered at 2pm and 12pm respectively. Our facility is on a 12 hour cycle, 6:30am-6:30pm. I've found that lower amounts of hormone work just as well, and sometimes better with C57s. Also I've found that using donors selected for body weight (13-15g at time of PMS injection) helps rather than using mice the same age and wildly different sizes. The number of blastocysts still varies day to day, but averages out over time. We inject 2-3 days per clone, and get plenty of chimeras. We flush blastocysts, and I wouldn't flush earlier than morulae--culture conditions can be tricky.

>>> Karin van Veen - 8/15/96 6:52 AM >>>

Hi, you all!

From kveen at nki.nl Tue Aug 20 11:17:30 1996 From: kveen at nki.nl (Karin van Veen)

Subject: blastocyst culture THANKS Message-ID:

Hello to everybody!

I want to thank everybody that reacted at my call for help concerning blastocyst-yield problems. Overall I can say that we are not doing things very wrong, most people use more or less the same techniques and time scedules. With some minor variations here and there. There where some tips I'm surely gonna try! I was glad to hear that almost everybody has or had the same problems as we have. Blastocyst yields from B6 mice varies a lot! It goes better and worse, without any clear reason. Last week I had a good week again, on three injection days, using 40 superovulated mice, I made 8 fosters, so almost 2 clones! I know, it is not fantastic, but I was happy. I'll let you know when I find THE answer to the problems. Bye, Karin. The Netherlands Cancer Institute Amsterdam

From tjf at uci.edu Tue Aug 20 10:28:50 1996 From: tjf at uci.edu (Tom Fielder)

Subject: Pricing Message-ID:

I am setting up a transgenic mouse core facility at UC-Irvine. We are trying to decide on our recharge rates, and I was hoping to get some feedback as to what other transgenic core facilities charge. We are planning to offer 2 levels of service. The basic package would be a fee per round of injection, with no guarantees as to outcome. The deluxe package would be a larger fee but would guarantee a certain number of transgenic pups (maybe 2?) (with perhaps a limit on the total number of tries). I'll try to extract as much info as I can from web pages, so if your prices are clearly posted (and readily accessible, e.g., through the Rat and Mouse Home Page, etc.), don't bother to respond. You can make it very brief, with just the price and what's guaranteed, and I will summarize the results for the list (minus names). I will post this same request on the Embryo Mail List and Compmed. Thanks!! Tom

From magree00 at pop.uky.edu Wed Aug 21 10:27:02 1996 From: magree00 at pop.uky.edu (Mike Green) Date: Wed Aug 20 11:52:52 2003 Subject: DNA quality/concentration Message-ID: <[email protected]>

When one of our researchers decides to produce transgenic mice, we provide a standard protocol for preparing the DNA for microinjection. In the past we've had a small number of labs who have produced generally high quality DNA for microinjection. Now that the number of facility users is increasing, we're implementing a pre-injection screening of the DNA to be injected. We plan to run a mini-gel to verify that there is a single band of the size and concentration claimed by the researcher.

Do other facilities do pre-injection screening? If so, what do you look for?

Would a spectrometer reading also be useful? Since most samples are of low volume and low concentration, I've been looking into the 7ul "ultra microvolume" cells for our GeneQuant. Does anyone have experience with these?

Mike Green University of Kentucky Transgenic Facility [email protected] 333 Combs phone: 606.257.2118 800 Rose St fax: 606.257.8940 Lexington, KY 40536 http://www.uky.edu/Transgenic/

From S.Tan at Anatomy.Unimelb.EDU.AU Thu Aug 22 14:17:47 1996 From: S.Tan at Anatomy.Unimelb.EDU.AU (Seong Seng Tan) Date: Wed Aug 20 11:52:52 2003 Subject: DNA quality/concentration Message-ID:

I would certainly recommend gel checking for both band size and DNA concentration. Lately I noticed that such purified DNA when stored at 4oC for a long time (over 12 months) are no longer able to produce transgenic mice by pronuclei injection. This is true even for DNA samples that have been proven to generated transgenic founders. Has anyone had the same experience?

>When one of our researchers decides to produce transgenic mice, we provide a >standard protocol for preparing the DNA for microinjection. In the past >we've had a small number of labs who have produced generally high quality >DNA for microinjection. Now that the number of facility users is increasing, >we're implementing a pre-injection screening of the DNA to be injected. We >plan to run a mini-gel to verify that there is a single band of the size and >concentration claimed by the researcher. > >Do other facilities do pre-injection screening? If so, what do you look for? > >Would a spectrometer reading also be useful? Since most samples are of low >volume and low concentration, I've been looking into the 7ul "ultra >microvolume" cells for our GeneQuant. Does anyone have experience with these? > > >

Seong-Seng Tan

======From pinkert at cmed.bhs.uab.edu Thu Aug 22 07:43:11 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert) Date: Wed Aug 20 11:52:52 2003 Subject: DNA quality/concentration Message-ID: <[email protected]>

> Date: Thu, 22 Aug 1996 13:17:47 +1100 > To: [email protected] > From: [email protected] (Seong Seng Tan) > Subject: Re: DNA quality/concentration > Reply-to: [email protected]

> I would certainly recommend gel checking for both band size and DNA > concentration. Lately I noticed that such purified DNA when stored at 4oC > for a long time (over 12 months) are no longer able to produce transgenic > mice by pronuclei injection. This is true even for DNA samples that have > been proven to generated transgenic founders. Has anyone had the same > experience? No we haven't seen an absolute result like this. What does the gel checking show you after a year? Is there degradation, additional bands, or just a concentration difference? Although we too have had doubts about long-term storage, we routinely store aliquots at -20 and we have used an aliquot of a 'test' gene over a two year period successfully. C.A. Pinkert

From Allison_F._Treloar at ccmail.bms.com Thu Aug 22 09:09:07 1996 From: Allison_F._Treloar at ccmail.bms.com (Allison F. Treloar) Date: Wed Aug 20 11:52:52 2003 Subject: DNA quality/concentration Message-ID: <[email protected]>

>>...Lately I noticed that such purified DNA when stored at 4oC for a long time (over 12 months) are no longer able to produce transgenic mice by pronuclei injection. This is true even for DNA samples that have been proven to generated transgenic founders. Has anyone had the same experience? ------

Absolutely! We routinely do a quick agarose check gel on stocks after 6 months to make sure they haven't degraded (noted as multiple bands or ladders). To keep our stocks fresh, we also keep an aliquot frozen just in case there is degredation in our working solution. (Don't freeze/thaw your stocks often, though!) Depending on your assay, you may still get positives, even with degraded samples, but they won't be expressors because the whole gene construct probably isn't intact.

In response to S-S Tan's questions: We have the investigator send us a gel picture of the construct band to show a single band is indeed present. We don't require a spec reading, but it's a nice addition. Have the investigator supply it! It sure is a lot easier to have them do it than to spend money buying yourself new equipment.

Allison Treloar Technical Supervisor - Transgenics Bristol-Myers Squibb [email protected]

"I try to take one day at a time, but sometimes several days attack me at once."

From kelly at citi2.fr Thu Aug 22 17:12:18 1996 From: kelly at citi2.fr (U344) Date: Wed Aug 20 11:52:52 2003 Subject: 129SV Peculiarities... Message-ID:

Hello, we've got K.O. mice which are 129/bl6 hybrids, and are trying to get our phenotypes back onto 129. Problem is, we're having a hard time getting these animals to reproduce... very few, pregnancies, then very low litter yields, or many pup deaths. Does anyone have experience with 129SV mice who could let me know if these peculiarities are common with this strain, or what I could do to make them happier? Any help would be greatly appreciated.

cheers...

'Man... I don't EVEN have an opinion...' From crussell at ResGen.COM Thu Aug 22 10:23:32 1996 From: crussell at ResGen.COM (Chris Russell) Date: Wed Aug 20 11:52:52 2003 Subject: 129SV Peculiarities... Message-ID: <[email protected]>

Some info from another mouse list:

We have backcrossed a 129 ES cells derived knock-out mice into B6. After 5 generations we ran into problems of very low fertility. We used good B6 partners for all mating, thus the problem is not B6.

Decrease in fertility is common in intercrosses during the production of congenic lines but are not supposed to happen in backcrosses. During the 5th and 6th generations of our backcross, we only obtained 1- 2 successiful litters of 4-6 animals after a year of mating each offspring with a healthy B6 partner.

Anyway we were able to survive the crisis, and after the 7th backcross generation the fertility problem disappeared.

Genetic heterogeniety effects in KO mice is getting more noticed. Thus it is wise to carefully plan one's mating strategies or one's choice of ES cells.

> Hello, we've got K.O. mice which are 129/bl6 hybrids, and are >trying to get our phenotypes back onto 129. Problem is, we're having a >hard time getting these animals to reproduce... very few, pregnancies, then >very low litter yields, or many pup deaths. Does anyone have experience >with 129SV mice who could let me know if these peculiarities are common >with this strain, or what I could do to make them happier? Any help would >be greatly appreciated. > > cheers... > >INSERM Unite 344 Endocrinologie Moleculaire >156 rue de Vaugirard, 75730 Paris cedex 15 France > >'Man... I don't EVEN have an opinion...' > > > > > > > *********************************************************************** **** Christopher G. Russell, Ph.D. RESEARCH GENETICS, INC Resources for Research -_+ _+_+ ++-_ _+-_ _+_ _++ _+-_ -+ -+ -+ +- +- -+ + -+ -+ -+ + -+ -+ -+ -+ +- -+ -+ -+ -+ -+ -+ -+ -+ -++-+ -+_-+ -+_+ -++- -+_++ -+_+ 2700 Memorial Parkway SW Huntsville, AL 35801 Phone 1-800-533-4363, ext 2211 or Direct 1-800-711-2089 U.K. 0-800-89-1393 Fax 1-205-551-1021 or 1-205-536-9016 [email protected] *********************************************************************** ******

From Allison_F._Treloar at ccmail.bms.com Thu Aug 22 12:04:43 1996 From: Allison_F._Treloar at ccmail.bms.com (Allison F. Treloar) Date: Wed Aug 20 11:52:52 2003 Subject: 129SV Peculiarities... Message-ID: <[email protected]>

>129/bl6 hybrids....very few, pregnancies, then very low litter yields, or many pup deaths .

129 females are not good mothers to start off with. All the problems you describe above are common to this strain. I don't know how to increase the pup yields (superovulation?), but you can combat the pup death issue by fostering off the pups to better ICR moms right away. We keep a few foster moms going for this purpose (about 2 foster litters drop/week). Couldn't hurt ;)

Allison Treloar Technical Supervisor-Transgenics Bristol-Myers Squibb Co. From tjf at uci.edu Thu Aug 22 17:33:29 1996 From: tjf at uci.edu (Tom Fielder) Date: Wed Aug 20 11:52:52 2003 Subject: Source for M2 medium Message-ID:

Help! Does anyone know of a source for M2 medium (for manipulation of mouse eggs) besides Sigma? They're back-ordered for 3-4 weeks. (Yes, I have the recipe, but not all the ingredients, nor the time to run quality control on home-made stuff). Tom

From browng at medicine.wustl.edu Fri Aug 23 07:58:44 1996 From: browng at medicine.wustl.edu (Gary Brown) Date: Wed Aug 20 11:52:52 2003 Subject: Source for M2 medium In-Reply-To: Message-ID:

I think that Gibco-BRL have M2, M16, BMOC (possibly more). Check out this source.

Enjoy,

On Thu, 22 Aug 1996, Tom Fielder wrote:

> Help! Does anyone know of a source for M2 medium (for manipulation of > mouse eggs) besides Sigma? They're back-ordered for 3-4 weeks. (Yes, I > have the recipe, but not all the ingredients, nor the time to run quality > control on home-made stuff). > Tom > > Thomas J. Fielder > UC-Irvine > University Lab Animal Resources > Transgenic Mouse Facility > 147 BSA > Irvine, CA 92697-1310 > e-mail: [email protected] > phone: 714-824-8579 > FAX: 714-824-2003 > > > > >

From Allison_F._Treloar at ccmail.bms.com Fri Aug 23 09:11:00 1996 From: Allison_F._Treloar at ccmail.bms.com (Allison F. Treloar) Date: Wed Aug 20 11:52:52 2003 Subject: Source for M2 medium Message-ID: <[email protected]>

>Help! Does anyone know of a source for M2 medium (for manipulation of mouse eggs) besides Sigma?

I will be one of the hundreds of replies you'll receive who will recommend Specialty Media, Inc. 800.543.6029

They supply M2 (and other common media) in liquid or lyophilized powder (my favorite because of the longer storage time).

liquid (50ml) MR-015-D $16.50 powder (5x10ml) MR-015P-5F $40.00

Good luck.

Allison

From mbo at avery.med.virginia.edu Fri Aug 23 09:26:07 1996 From: mbo at avery.med.virginia.edu (M. B. Ober) Date: Wed Aug 20 11:52:52 2003 Subject: Source for M2 medium Message-ID:

I get my M2/M16 from Specialty Media, 1-800-543-6029. They have ready-to-thaw and rehydratable versions.

Maggie Ober Transgenic Mouse Core Facility University of Virginia

From tjf at uci.edu Fri Aug 23 09:53:11 1996 From: tjf at uci.edu (Tom Fielder) Date: Wed Aug 20 11:52:52 2003 Subject: Source for M2 medium Message-ID: <[email protected]>

Gary, I had already checked the Gibco catalog and didn't see any of these (although I didn't call). However, 4 other people have already recommended Specialty Media, Inc. in New Jersey (800-543-6029). Fortunately, they have it in stock, although at more than twice the cost of Sigma. Tom

>I think that Gibco-BRL have M2, M16, BMOC (possibly more). Check out this >source. > >Enjoy, > >Gary Brown >E-mail: [email protected] or [email protected] >Web: http://ourworld.compuserve.com/homepages/TheBroons/ > "The Microinjection Workshop" > > >On Thu, 22 Aug 1996, Tom Fielder wrote: > >> Help! Does anyone know of a source for M2 medium (for manipulation of >> mouse eggs) besides Sigma? They're back-ordered for 3-4 weeks. (Yes, I >> have the recipe, but not all the ingredients, nor the time to run quality >> control on home-made stuff). >> Tom >> >> Thomas J. Fielder >> UC-Irvine >> University Lab Animal Resources >> Transgenic Mouse Facility >> 147 BSA >> Irvine, CA 92697-1310 >> e-mail: [email protected] >> phone: 714-824-8579 >> FAX: 714-824-2003 >> >> >> >> >> > > > > > Thomas J. Fielder UC-Irvine University Lab Animal Resources 147 BSA Irvine, CA 92697-1310 [email protected]

From mbo at avery.med.virginia.edu Mon Aug 26 12:45:23 1996 From: mbo at avery.med.virginia.edu (M. B. Ober) Date: Wed Aug 20 11:52:52 2003 Subject: PMS Gonadotropin Message-ID:

Sigma has just informed me that their Pregnant Mares' Serum Gonadotropin is on backorder. Last time they did this it was almost 6 months before they had it available. What is a good alternative source?

I've been using the 50 IU/vial lyophyllized PMSG. Is this overkill? When Sigma had their 50IU/vial hCG on backorder I took a chance on ICN's hCG (5000 IU for the price of one Sigma vial) and it seems to be working fine. Can I get away with cheaper PMSG also?

From sp3i at avery.med.virginia.edu Mon Aug 26 12:58:41 1996 From: sp3i at avery.med.virginia.edu ([email protected]) Date: Wed Aug 20 11:52:52 2003 Subject: PMS Gonadotropin Message-ID:

M >I've been using the 50 IU/vial lyophyllized PMSG. Is this overkill? When >Sigma had their 50IU/vial hCG on backorder I took a chance on ICN's hCG >(5000 IU for the price of one Sigma vial) and it seems to be working fine. >Can I get away with cheaper PMSG also?

It seems there's nothing to lose by trying the cheaper PMSG if you can get/find it.

Is it the case that 22 out of 28 hostesses were pregnant in the SM22 series? That's my read from the database. S

------Sonia Pearson-White, Ph.D. 804-982-0756 tel., 804-982-3993 fax. University of Virginia Medical Center, MR4 Room 3002, 300 Park Place Charlottesville, VA 22908

From PARLOW at AFP76.HUMC.EDU Mon Aug 26 10:30:50 1996 From: PARLOW at AFP76.HUMC.EDU (phone(310)222-3537 fax222-3432) Date: Wed Aug 20 11:52:52 2003 Subject: PMS GONADOTROPIN AVAILABLE Message-ID: <[email protected]>

REGARDING SUPPLY OF PMSG, TWO CONCERNS ARE

1) ACCURACY OF THE STANDARDIZATION 2) COST

PMSG IS CURRENTLY AVAILABLE FROM MY LABORATORY IN A RELIABLY STANDARDIZED FORMAT, IN ABUNDANT SUPPLY.

REQUESTS MAY BE SUBMITTED TO MY E-MAIL ADDRESS OR TO MY FAX LISTED ABOVE OR BY MAIL TO

DR. A. F. PARLOW HARBOR-UCLA MEDICAL CENTER PITUITARY HORMONES/ANTISERA CENTER 1000 WEST CARSON ST. TORRANCE, CA 90509

I WILL COME TO YOUR RESCUE.

/aFp ======your inquiry appears below ======

From: IN%"[email protected]" 26-AUG-1996 08:47:25.05 To: IN%"[email protected]" CC: Subj: PMS Gonadotropin

From jparkert at magnus.acs.ohio-state.edu Mon Aug 26 15:55:58 1996 From: jparkert at magnus.acs.ohio-state.edu (Janice Parker-Thornburg) Date: Wed Aug 20 11:52:52 2003 Subject: DNA quality/concentration Message-ID: <[email protected]>

I guarantee a set number of transgenics with each injection, and have found the DNA prep to be critical. Thus, rather than trust some of the researchers, I request a restriction digest along with a picture proving it is digested, and where the band of interest is circled. Then, I prep the DNA myself. I have had very good luck with this. No, I would not trust many researchers to prepare high quality DNA--especially if they have never done a microinjection before. As a molecular biologist myself, I know that one is often less than careful when prepping DNA for cloning and digests. However, as an injector, I also know how pure the DNA must be for this type of work. If you can, do it yourself. I can send you an easy prep that gives great DNA.

Jan Parker-Thornburg, Manager Transgenic Animal Facility The Ohio State University [email protected]

From h-marsha at nimr.mrc.ac.uk Tue Aug 27 10:42:39 1996 From: h-marsha at nimr.mrc.ac.uk (Heather) Date: Wed Aug 20 11:52:52 2003 Subject: PMS Gonadotropin Message-ID:

>Sigma has just informed me that their Pregnant Mares' Serum Gonadotropin is >on backorder. Last time they did this it was almost 6 months before they >had it available. What is a good alternative source? > >I've been using the 50 IU/vial lyophyllized PMSG. Is this overkill? When >Sigma had their 50IU/vial hCG on backorder I took a chance on ICN's hCG >(5000 IU for the price of one Sigma vial) and it seems to be working fine. >Can I get away with cheaper PMSG also? > >Maggie Ober >Transgenic Mouse Core Facility >University of Virginia

Hello Maggie,

This may not help you much as it involves shipping costs, but if you're really stuck, then Intervet Labs in Cambridge, UK. may be able to help. They provide a box of 5 vials PMS (1000IU/vial) for ?17.40. This works out at about ?3.50 per 1000 IU, where Sigma charge ?33.70 for the same amount!! Intervet's hCG is also supplied at 5 vials per box, but at 1500IU per vial (?38 per box). I can give you their details if you want to take this further.

Heather Marshall Division of Developmental Neurobiology National Institute for Medical Research The Ridgeway Mill Hill London. NW7 1AA Tel:(0181-959-3666) [email protected]

From jparkert at magnus.acs.ohio-state.edu Thu Aug 29 16:58:53 1996 From: jparkert at magnus.acs.ohio-state.edu (Janice Parker-Thornburg) Date: Wed Aug 20 11:52:52 2003 Subject: B-cell specific expression Message-ID: <[email protected]>

This is just a quick query to see if I can get a response here prior to doing some library legwork. I am working with an investigator who wants to have his gene expressed in B-cells in transgenic mice. I was wondering if anyone out there either has, or knows of someone who has a plasmid containing the mouse IgG enhancer with an appropriate promoter and subsequent cloning sites that has worked well in transgenics. The one he has now has an IgG enhancer of unknown origin followed by the promoter for a housekeeping gene. I'm rather skeptical that it will work to the extent that he needs it. I appreciate any leads.

Thanks much.

Jan Parker-Thornburg

From Dnxrh at aol.com Thu Aug 29 17:35:53 1996 From: Dnxrh at aol.com ([email protected]) Date: Wed Aug 20 11:52:52 2003 Subject: B-cell specific expression Message-ID: <[email protected]>

You can do an on-line keyword search (without leaving your seat) of the Oakridge National Laboratories Transgenic and Targeted Mouse Database at: http://www.ornl.gov/TechResources/Trans/hmepg.html

One suggestion. Once you have identified potential promotors by doing a "B-Cell" keyword search, you might want to go back and use the promotor itself as a keyword for a follow up search. You never know what other tissues a particular promotor might also express in and that can be relevant.

Rick Huntress DNX Transgenics [email protected] 508-779-0189 (phone) 508-779-0190 (fax)

My opinions are my own and not necessarily that of my employer.

From Allison_F._Treloar at ccmail.bms.com Fri Aug 2 21:50:53 1996 From: Allison_F._Treloar at ccmail.bms.com (Allison F. Treloar)

Subject: Transgenic Barriers Message-ID: <[email protected]>

Hi folks,

[email protected]

From mgallardo at wsu.edu Fri Aug 2 21:36:49 1996 From: mgallardo at wsu.edu (Mike Gallardo)

Subject: Transgenic Barriers Message-ID: <[email protected]> At 03:50 PM 8/2/96 -0500, you wrote: >Hi folks, > >I am trying to get a feel for how many institutions are using barrier conditions >for their transgenic/knockout colonies. Please reply to me off list and I'll >sumarize for the group. Don't forget to include where you're from! > >no/low barrier = conventional caging and access (+/- filter tops on caging) > >modified barrier = access restrictions, gowning, isolators, (+/- autoclaving) > >full barrier = shower in, isolator caging, autoclaved food and bedding > >Thanks in advance! >Allison Treloar >Technical Supervisor - Transgenics >Bristol-Myers Squibb > >[email protected] > > >Here at Wegner Hall Vivarium/WSU we use isolator caging, autoclaved food and bedding. We do not shower in, we use steril gowns, booties, mask, hair nets and gloves. > >

(''`-''-/").___..--''"`-._ `o_ o ) `-. ( ).`-.__.`) (_Y_.)' ._ ) `._ `. ``-..-' _..`--'_..- / /--' _. .' (il).-'' (li).' ((!.-' Mike Gallardo BS MS LAT Administative Manager

GO COUGS!!!! Wegner Hall Vivarium G-44 Washington State University Pullman, Wa 99164-6510 Phone (509)335-0977 Fax (509)335-0162 e-mail [email protected] "Brains are for thinking--pencils are for remembering"

From fmargoli at umabnet.ab.umd.edu Fri Aug 2 23:13:51 1996 From: fmargoli at umabnet.ab.umd.edu (Frank L. Margolis)

Subject: No subject Message-ID: Anyone-

Is there an up-to-date master list of knockout and/or transgenics that is maintained anywhere?

Thanks

From ignelzi at umich.edu Fri Aug 2 22:15:36 1996 From: ignelzi at umich.edu (Michael A. Ignelzi, Jr.)

Modified barrier, access restrictions, gowning, isolators, (+/- autoclaving), Univ of Michigan School of Dentistry (transgenic and knockout colonies)

>Hi folks, > >I am trying to get a feel for how many institutions are using barrier >conditions >for their transgenic/knockout colonies. Please reply to me off list and I'll >sumarize for the group. Don't forget to include where you're from! > >no/low barrier = conventional caging and access (+/- filter tops on caging) > >modified barrier = access restrictions, gowning, isolators, (+/- autoclaving) > >full barrier = shower in, isolator caging, autoclaved food and bedding > >Thanks in advance! >Allison Treloar >Technical Supervisor - Transgenics >Bristol-Myers Squibb > >[email protected] >

From UCIVET at uci.edu Fri Aug 2 23:21:06 1996 From: UCIVET at uci.edu (Clifford R ROBERTS)

Subject: Transgenic Barriers Message-ID: <[email protected]> low barrier: microisolator cages, changed in Class II hood, Masks, lab coats, gloves.

Cliff Roberts Univ Calif, Irvine ______Subject: Transgenic Barriers From: [email protected] at biosmtp Date: 8/2/96 1:19 PM

Hi folks,

[email protected]

From pinkert at cmed.bhs.uab.edu Fri Aug 2 20:17:28 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert)

Subject: Transgenic Barriers Message-ID: <19960803002112601.AAD148@cap1>

> Date: Fri, 02 Aug 96 14:21:06 PST > To: [email protected] > Subject: Re: Transgenic Barriers > Reply-to: [email protected] After pulling up a number of responses to the barrier question - with each individual describing a facility, kindly respond directly to Allison Treloar ([email protected]) as it is getting to be a bit much.

From p.sobieszczuk at ic.ac.uk Sat Aug 3 15:53:41 1996 From: p.sobieszczuk at ic.ac.uk (Peter Sobieszczuk)

Subject: "TgList Backstage" (admin. news *1) Message-ID:

Dear Subscribers,

Therefore, an explanation: if you use "reply" function of your e-mail package (e.g. "reply" from "message" menu in Eudora) remember it will go back to the list, not only to the original sender. To send your reply (or any message) just to one person use "new" message or type that address in the "to:" field yourself. It would also appear to be sensible to cut out everything what is not relevant from the copy of the original letter in your "reply to the list" message.

Thank you

Thanks,

Allison

Technical Supervisor-Transgenics Bristol-Myers Squibb Co. [email protected] ______From MEISLERM at hg-basic1mail.hg.med.umich.edu Sat Aug 3 19:04:43 1996 From: MEISLERM at hg-basic1mail.hg.med.umich.edu (MEISLERM)

Subject: brain and neuron specific promoters Message-ID:

Does anyone know of a recent review of promoters with in vivo specificity for subsets of neurons, glia, or brain regions? We are looking for promoters specific for brain regions, spinal cord, motor neurons, etc. Thanks for your suggestions.

Miriam Meisler, Ph. D. Department of Human Genetics University of Michigan 4708 Medical Sciences II Ann Arbor, MI 48109-0618 tel 313 763 5546; FAX 313 763 9691.email: [email protected]

From gstuart at UVic.CA Sun Aug 4 01:01:42 1996 From: gstuart at UVic.CA (Greg Stuart)

Subject: Rat liver anatomy question Message-ID: <[email protected]>

Hello - I'd like to draw on the collective experience of the subscribers. Can anybody describe where the following regions of the rat liver are located?

- centrilobular region - midzonal hepatocytes - periportal regions

Also, are these regions found in each lobe of the liver, or are they specific to particular lobes or regions?

Please respond directly, thanks. Cheers, Greg :)

From hage at med.unc.edu Sun Aug 4 03:01:14 1996 From: hage at med.unc.edu (John Hagaman)

>Hi folks, > >I am trying to get a feel for how many institutions are using barrier >conditions >for their transgenic/knockout colonies. Please reply to me off list and I'll >sumarize for the group. Don't forget to include where you're from! we us modified barrier = access restrictions, gowning, isolators, (+/- autoclaving, both )

> >[email protected] >

966-6912 office 966-8800 Fax

From kelly at citi2.fr Mon Aug 5 23:12:58 1996 From: kelly at citi2.fr (U344)

Subject: mouse testosterone levels Message-ID:

Hello, does anyone know of a way to measure mouse testosterone levels, or of anyone who is? I'd really like to avoid having to set up an ELISA for a few samples... cheers...

From browng at medicine.wustl.edu Mon Aug 5 13:53:29 1996 From: browng at medicine.wustl.edu (Gary Brown)

Subject: WWW resource Message-ID:

Just a brief wee message here from St Louis. At the risk of tooting my own horn I'd like to let anyone interested know about my microinjection / transgenisis related homepage. Apologies go to those who may have seen this on the COMPMED listings, or to those who subscribe to this list after my listing THIS list on the page in question! Anyways, here it is...

Page Name: The Microinjection Workshop Page address: http://ourworld.compuserve.com/homepages/TheBroons/homepage.htm How best to find: Although I have registered with most of the widely used search engines, Yahoo is still a problem.. Best to use the Starting Point (www.stpt.com) and select the combination Metacrawler engine searching on "microinjection".

I'm trying to build up an E-mail mailing list to alert when the page is updated (approx. 1 x per month). If interested please reply OFF-LIST tothe following:

[email protected]

Thanx :-)

From Allison_F._Treloar at ccmail.bms.com Mon Aug 5 15:59:37 1996 From: Allison_F._Treloar at ccmail.bms.com (Allison F. Treloar)

Subject: Summary: Tg Barriers Message-ID: <[email protected]> First and foremost, a big thanks to everyone who responded to my question regarding barrier conditions for trangenic colonies. Here's the outcome:

17 respondents 3 facilties use low/no barrier conditions 10 facilities use modified barrier conditions 4 facilities use full barrier conditions low/no barrier respondents stated that these conditions were used primarily for single transgenic lines or areas where investigators could generally access transgenic stock animals. modified barrier users seem to rely on access restrictions, gowing, dedicated staff, isolator caging and autoclaved food/bedding, but don't go as far as shower-in or sterile clothing/scrubs. Most cage changing/animal manipulation is done in laminar flow hoods. This is certainly the most popular, flexible and cost effective option based on current opinion. full barriers were used by a few facilities and included air showers, sterile clothing/scub requirements as well as all the modified barrer conditions. No respondents with full barriers used shower-in.

Thanks, again for all your help.

Allison

[email protected]

From gina_seeburger at Merck.Com Mon Aug 5 20:07:08 1996 From: gina_seeburger at Merck.Com (Gina Seeburger)

Subject: "TgList Backstage" (admi Message-ID: <199608052001.QAA29238@igw2>

RE>"TgList Backstage" (admin.

------Date: 8/3/96 10:56 AM To: Gina Seeburger From: [email protected] Dear Subscribers,

Copies of the first two posting from the last week: ______Hello Transgenic List Subscribers: Can anyone direct me to any studies, papers, or general information, regarding stress indicators of mice that have been placed in the dark or light reduced areas for 24 hours or longer ?

Thank you

Thanks,

Allison

Technical Supervisor-Transgenics Bristol-Myers Squibb Co. [email protected] ______

From TSAUNDER at hg-basic1mail.hg.med.umich.edu Tue Aug 6 15:57:55 1996 From: TSAUNDER at hg-basic1mail.hg.med.umich.edu (TSAUNDER)

Subject: Meeting Announcement Message-ID:

THE UNIVERSITY OF MICHIGAN CENTER FOR ORGANOGENESIS 1st International Symposium

Molecular Control of Organogenesis Saturday, October 5, 1996 Horace H. Rackham Graduate School Amphitheater The University of Michigan Ann Arbor, Michigan

8:45-9:00 Opening Remarks, Dr. Giles G. Bole, Dean, The University of Michigan Medical School Dean

9:00-9:50 Dr. Eric Olson, University of Texas, Dallas, Texas USA, "Genetic control of myogenesis"

9:50-10:40 Dr. Peter Gruss, Max Planck Institute of Biophysical Chemistry, Germany, "Genes required for the development of the visual system"

10:40-10:50 Break

10:50-11:40 Dr. Cliff Tabin, Harvard Medical School, Boston, Massachusetts, USA, "Signals patterning the vertebrate limb"

11:40-12:30 Dr. Yuh Nung Jan, University of California, San Francisco, California, USA, "Molecular control of neural development"

12:30-2:30 Poster Session (lunch provided)

2:30-3:20 Dr. Paul Sternberg, California Institute of Technology, Pasadena, California, USA, "Precise formation of a hole: the C. elegans vulva"

3:20-4:10 Dr. Richard M. Harland, University of California, Berkeley, California, USA, "Re-use of signaling molecules in vertebrate axis formation and organogenesis"

4:10-4:20 Closing Remarks, Dr. Seigo Izumo, Acting Director, Center for Organogenesis, The University of Michigan

Friday, October 4, 1996 5:30-7:00 p.m.Meet the Speakers at an Open Reception, Museum of Art, The University of Michigan

For further information, please contact Michelle Shukait at 936-2499 or [email protected]

From hage at med.unc.edu Wed Aug 7 04:28:28 1996 From: hage at med.unc.edu (John Hagaman) Subject: White noise for Mouse Embryo Yield Message-ID:

Does anyone have experience re: noise in mouse rooms and how it may affect the yield of embryos? Do mice "get used to certain noises" Does with drawal or introduction of noise change the yield?

Responses off list will be summarized.

Thanks

966-6912 office 966-8800 Fax

From fmargoli at umabnet.ab.umd.edu Wed Aug 7 15:38:50 1996 From: fmargoli at umabnet.ab.umd.edu (Frank L. Margolis)

Subject: Knock out master list Message-ID:

Thanks to all for your responses to my query, they were most helpful.

Frank Margolis

From r-carver at nimr.mrc.ac.uk Thu Aug 8 17:47:28 1996 From: r-carver at nimr.mrc.ac.uk (Rick Carver)

Subject: LASA Transgenics Meeting Message-ID: <[email protected]> Dear All

I am now able to give details of the speakers for the forthcoming LASA Transgenics meeting.

Transgenesis- setting the scene Dr Roy Forster, Transgene Transgenic Animals in Developmental Biology Dr Reter Rigby, NIMR Transgenic Animals in Immunology Dr Mike Owen, ICRF Transgenic Animals in Production Proteins Dr Ron James, Pharmaceutical Proteins Ltd Transgenic Animals in Neuroscience Prof Rick Lathe, Univ of Edinburgh Transgenic Animals in Oncology Dr Frances Balkwell, ICRF Transgenic Animals in Pharmaceuticals Dr John McNeish, Pfizer US Discovery and Development Ethical and Legal Aspects of the Use of Dr Kenneth Boyd, Univ of Edinburgh Transgenic Animals

The cost will be 55 pounds stirling for LASA members and 75 pounds stirling for non-members. Registration forms are available from: LASA Speciality Sections, PO Box 3993, Tamworth, Staffs, B78 3QU, UK. The closing date for registrations is 23 Aug 96.

From gac at po.mri.montana.edu Thu Aug 8 00:36:49 1996 From: gac at po.mri.montana.edu (George Carlson)

Subject: discontinuation of strains Message-ID:

Most of the intra-H-2 recombinant B10 congenic strains developed by the late Dr. Jack Stimpfling of McLaughlin Research Institute will be discontinued as live stock in the near future. Following Dr. Stimpfling's retirement, a grant from the American Cancer Society allowed us to complete isolation of his more recently identified recombinant chromosomes, to refine localization of the crossover sites within H-2, to delimit the congenic intervals for each strain, and to preserve the strains for future use. We are now in the final stage of this project and expect to have all strains frozen before January 1997. DNA samples will continue to be available.

The strains are listed in: Turner et al. Meiotic recombination within the H-2K--H-2D interval: characterization of a panel of congenic mice, including 12 new strains, using DNA markers. Immunogenetics 1993, 38:332-340.

Additional data on our congenic panel are being prepared for publication.

Breeding pairs are available for any investigator who requests them. We only ask that investigators who "adopt" a strain make it available to others in the community and provide notification if they stop the line.

I will provide a listing of the strains that we will continue as live stock after we get a better idea of interest. Please let me know if you need any of these mice. Suggestions of other appropriate lists for posting this notice would also be appreciated.

Thank you.

George A. Carlson ([email protected]) McLaughlin Research Institute 1520 23rd Street South Great Falls, Montana 59405 406 452 6208, fax: 454 6019 or 454 6004

We are currently undergoing major building renovations and this week alone, three of our litters have been eaten by their mothers. We suspect loud sudden noises are responsible.

Seong-Seng Tan

======

From fmargoli at umabnet.ab.umd.edu Thu Aug 8 16:11:34 1996 From: fmargoli at umabnet.ab.umd.edu (Frank L. Margolis)

Subject: Lists Message-ID:

To all-

In response to several requests I am posting a summary of all the info I have received about the availability of lists of Knockouts and transgenics and specific strains (without any editorial evaluation).

BioMedNet has at least a partial list of knockouts

JAX homepage is http://www.jax.org

Current list of induced mutant strains accepted for inclusion in the Induced Mutant Resource (IMR) at JAX is on the IMR home page- http://lena.jax.org/resources/documents/imr/

An index of strains is available at- http://lena.jax.org/resources/documents/imr/imr.html and more details at- http://lena.jax.org/resources/documents/imr/imrdata.html for queries about any JAX mice contact- [email protected]

Also the following web sites have info that may prove to be useful- NIH transgenic directory Tbase http://www.gdb.org/Dan/tbase/tbase.html

Mouse and rat research home page- http://www.cco.caltech.edu/~mercer/htmls/rodent_page.html#homepages and as was also pointed out to me: 1-a Medline search for info on specific genes or strains as well as 2- right here where I started all this.

Thanks again for all your input.

Frank Margolis

From gstuart at UVic.CA Thu Aug 8 15:37:43 1996 From: gstuart at UVic.CA (Greg Stuart)

Subject: Isolation of bone marrow & colon tissues Message-ID: <[email protected]>

Hello everyone. Does anybody have protocols that they could share for the isolation of rat (or rodent) bone marrow and colon tissue? Thanks, Greg :)

From CLUCK.RICHARD_S+ at LEXINGTON.VA.GOV Thu Aug 8 21:45:00 1996 From: CLUCK.RICHARD_S+ at LEXINGTON.VA.GOV (CLUCK.RICHARD_S+)

Subject: Copy of: Re: White noise for Mouse Embryo Yield Message-ID: <[email protected]>

You wrote:

While working at the University of Kentucky, I found a number of interuptions in breeding colonies due to the noise caused by construction. Liters were either canabalized by the mother or breeding stopped completely. Hope this helps.

------( )_( ) Richard S. Cluck, B.S., LATG o o Supervisor, VMU, VA, Lexington ==\o/== Research 151 /\ ) Cooper Drive / \ ( Lexington, KY 40511 / \) Phone: 606-281-4927 Fax: 606-281-4989 E-Mail: cluck,[email protected] ------

From r-lovell at nimr.mrc.ac.uk Fri Aug 9 17:38:45 1996 From: r-lovell at nimr.mrc.ac.uk (Robin Lovell-Badge)

Subject: Copy of: Re: White noise for Mouse Embryo Yield Message-ID:

Dear All

There is considerable published and anecdotal stuff on noise in animal rooms. Excessive noise, e.g. from building works, can have all sorts of effects which will lower reproductive efficiency. Mice seem to particularly dislike sudden noises, so it can be beneficial to have music playing (classical or modern, but not too much rap !) during the day when cages are being changed, etc. But again, it should not be very loud, just enough to mask the dropping of cages, screams of students, etc.

If anyone is desperate I can try to locate published work on this - but it is not recent and may take a while.

Robin Lovell-badge >You wrote: > >>Does anyone have experience re: noise in mouse rooms and how it may affect >>the yield of embryos? Do mice "get used to certain noises" Does with >>drawal or introduction of noise change the yield? >> > > While working at the University of Kentucky, I found a number of >interuptions in breeding colonies due to the noise caused by construction. >Liters were either canabalized by the mother or breeding stopped >completely. Hope this helps. > > >------> ( )_( ) > Richard S. Cluck, B.S., LATG o o > Supervisor, VMU, VA, Lexington ==\o/== > Research 151 /\ ) > Cooper Drive / \ ( > Lexington, KY 40511 / \) > Phone: 606-281-4927 > Fax: 606-281-4989 > E-Mail: cluck,[email protected] >------

From dknudsen at scruznet.com Tue Aug 13 04:32:45 1996 From: dknudsen at scruznet.com (Dave Knudsen)

Charmaine -

Dave Knudsen DVM, MS, DACLAM Scotts Valley, California, USA *** "In my judgement it is either an enigma or some kind of bug. If it dies, I will take it apart and see what its arrangements are. I never had a thing perplex me so." Mark Twain, 1893 (from "Extracts from Adam's Diary") ***

Many thanks.

David Bowtell.

Lab 61-3-96561287 Office 61-3-96561296 Fax 61-3-96561411 email: [email protected] From wixson at biovax.dnet.basf-ag.de Tue Aug 13 18:16:55 1996 From: wixson at biovax.dnet.basf-ag.de (Sally Wixson, VMD, MS)

Many thanks for the advice, Sally K, Wixson, VMD, MS From wixson at biovax.dnet.basf-ag.de Tue Aug 13 18:16:55 1996 From: wixson at biovax.dnet.basf-ag.de (Sally Wixson, VMD, MS)

Many thanks.

David Bowtell.

From dknudsen at scruznet.com Tue Aug 13 04:32:45 1996 From: dknudsen at scruznet.com (Dave Knudsen)

Charmaine -

From browng at medicine.wustl.edu Wed Aug 14 13:52:59 1996 From: browng at medicine.wustl.edu (Gary Brown)

Subject: Knockout advice In-Reply-To: <[email protected]> Message-ID:

Just throwing in my 2 cents worth...

On Tue, 13 Aug 1996, Sally Wixson, VMD, MS wrote:

> .....what strain of vactomized males do you use and why? How do you assess > their sterility (other than ordering then vasec. by the vendor)? How often do > you use vasec. males? (my invesitagors use each male once/week or less!)

I use Swiss Webster VAS-X males, ordered in from Taconic Farms. In my previous position we did our own vasectomies on C57Bl/6J males (pronuclear microinjection only) and assessed their sterility by co-housing with Swiss Webster (or other inexpensive white mouse strain) females (2) for approx. 6 weeks prior to use. Males were vasectomized at 4-5 weeks of age, excising a minimum of 1cm of vas deferens per side. > > .....what strain of embryo recipients do you use for ES cell work? Our donors > are C57BL/6, recipients are B6D2F1's. My team wants a "big" target so they keep > the females here three months or so on a high fat diet so they are big enough > to inject. Obviously, this leads to a waste of cage space. Many people use > ICR's or CD1's as recipients because they are a big target and good moms!

I have used B6D2F1s for recipients although now use Swiss Webster (occasionally ICR) females as they are a relatively large mouse strain. As for feeding them on a high fat diet I personally do not subscribe to the 'big target' philosophy - at least no bigger than using regular mouse chow for my chosen strain. Since we have a pool of SW females from which to pick those in estrus and individuals may be passed over several times, high fat diet pushes some of them to a size where surgery becomes more difficult due to the increased fat deposits around the oviduct/ovary/UT junction, and I believe that the increased incision size in the body wall that this necessitates increases the risk to the animal. > > .....in breeding chimeras, our PI insists that each chimera must produce100 > pups (black) before being considered a failure at germ line transmission. Many > of the chimeras are lowly sterile or sterile, so this format entails breeding > formany months to more than a year. Several other labs told me their chimeras > get three matings and if no agouti pups are produced, they are ENA'd.

This is insane! My initial reaction would be to tell your PI to get a life! Our policy is to discard all female chimeras (low germline transmission, increased sterility in the F1 generation when successful) and to discard all >50% (by color coat) male chimeras. 3 matings (strikes) and you're out sounds appropriate to me. > > .....do you produce your own ES cells or get them from donation/purchase from > other labs? We are currently buying the RW4 ES cell line from Genome Systems. > Do you require MAP testing of the new ES cell lines?

Our lab has approached other labs for ES cells, although we are trying to establish our own cell line currently. With reference to the RW4 cell line, I have used this effectively on several occasions and note that it has a high color coat conversion rate. I also know the individual who created this cell line, and can forward any questions you may have to her. Our institution requires MAP testing of all ES cell lines for use in mice. > > .....do you superovulate your females for embryo donation? My PI says this doe > not yield viable embryos for ES cell work, so they breed 4-5X the number of > females they rpedict will be needed for each injectionso that they have enough > embryos. > Yes, I superovulate females for embryo donation. I typically superovulate 7 C57Bl/6J females for one day's effort, with a variation between 20-25 morula per plugged female. I culture these in the incubator overnight to blastocysts with an 85-95% conversion rate. The females used are between 3.5-4.5 weeks old. I use Sigma Chemical Co. new embryo tested PMSG and HCG. My stud males are mated no more than once weekly. There is a reference for superovulating females for blastocyst production, but I would have to look it up on Medline. Sounds to me like your PI needs to do some homework!

Hope this helps!

From kveen at nki.nl Thu Aug 15 11:52:09 1996 From: kveen at nki.nl (Karin van Veen)

Subject: blastocyst culture Message-ID:

Hi, you all!

From browng at medicine.wustl.edu Thu Aug 15 13:23:30 1996 From: browng at medicine.wustl.edu (Gary Brown)

Subject: blastocyst culture In-Reply-To: Message-ID:

On Thu, 15 Aug 1996, Karin van Veen wrote: > Can someone give me some tips concerning the time schedule on the > superovulation. In what time intervals do you inject the hormones, and > isolate the embryo's? > We use C57/bl6 mice as donors.

We use a 12hr/12hr light-dark cycle from 4am to 4pm in our facility, coupled with 47hrs between PMSG and HCG. These hormones are typically given at 10.30am and (47hrs later) 9.30am repectively for this light cycle. We use hormones obtained through Sigma Chemical Co. (Embryo culture tested - new product) and have been obtaining yields of MORULAE at a rate of 20-25 per plugged C57Bl/6J female. These morulae are cultured overnight to blastocysts for ES cell injection in Gibco's modified BMOC-3 culture media with pen/strep.added in a 37deg C 5% CO2 incubator.

> We are also thinking, when it should be a flushing problem, to isolate the > zygote on day 0,5, and culture them into blastocyst to be injected. Has > anyone experience on this? How long do you have to culture the zygotes to get > blastocysts, are they good to be injected with ES cells? > All tips are welcome! > For culture past day 0.5, you would need 3 more days and some consistently good media. I suggest you flush morulae! Although I doubt that flushing would present too much of a problem, this is how I do it. Using a short bevel stainless 30g hypo., introduce the needle at the last oviduct loop before the UT junction and flush media through this area expelling the embryos out of the uterus (cut 1cm down from the oviduct). If this is problematical, flush through the infundibulum (requires a little more finesse).

Hope this helps out! Gary Brown Email: [email protected] or [email protected] Web: http://ourworld.compuserve.com/homepages/TheBroons/ "The Microinjection Workshop"

Test - please ignore

Subject: Test please ignore Message-ID:

From gina_seeburger at Merck.Com Thu Aug 15 21:15:46 1996 From: gina_seeburger at Merck.Com (Gina Seeburger)

Subject: blastocyst culture Message-ID: <[email protected]>

REPLY o REPLY o REPLY o REPLY RE>blastocyst culture Karin, We were having problems too here in the US, but giving the first injection(PMSG, Sigma brand) between 3:00pm and 4 (.3cc, .7.5IU) and the second (hCG) around 1:00 seems to help. We are also using C57BL/6 donors for blast injections. Collected around 7:30am. Also summer seems to create a low. Good luck Gina Seeburger Merck & Co,Inc. [email protected]

------Date: 8/15/96 5:57 AM To: Gina Seeburger From: [email protected]

Hi, you all!

Subject: oocyte injection Message-ID:

I am brand new to the field of transgenics, and I am setting up a transgenic core facility at UC-Irvine. I am still trying to optimize my pipets for injection of DNA into oocytes. I'm using a Sutter P-97 puller, and 1mm OD x 0.78mm ID capillaries with internal filament. I got some pipets that seemed to work very well yesterday, judging by the swelling of the pronucleus and no visible cell contents leaking out after withdrawing the pipet, but this morning 65% of the injected cells had died (cytoplasm was condensed and dark brown), while none of the uninjected controls looked like this. I'm afraid that the taper of the needle is too abrupt near the tip (i.e., it gets too big too fast). A very patient person at Sutter is helping me with the pipets, but my question for the list is this: Is the condensed, dark brown cytoplasm indicative of mechanical trauma, or is it a result of bad DNA/buffer, or something else? A second question: The books recommend using females that are 24-25 days old for superovulation. What happens if you use older ones? Will the yield of oocytes be significantly less? I've been using 6-8 week old FVB mice and getting about 30 eggs/mouse, but I've noticed that a large number of them aren't fertilized, even from the females that were obviously plugged. Any thoughts? Tom

From MKENNEDY at chmeds.ac.nz Sat Aug 17 11:12:42 1996 From: MKENNEDY at chmeds.ac.nz ([email protected])

Subject: Double KO with increased G418 Message-ID: <[email protected]>

Date sent: 17-AUG-1996 22:06:32

Hi Dave,

Just got back from this year's Queenstown Meeting, which (for my sins) I organised.

>We have used increased G418 concentration as a means of selecting homozgous >cells on two occassions. One construct had a pGKNeo casssette and the cells >were still laughing in 4mg/ml (we blinked first). The second construct was >promoterless, the cells were marginally G418 resistant and it worked like a >dream. The paper by Mortensen (MCB 12, 2391) used a pGKNeo cassette but I >am unclear whether it was the point mutant (low activity) neo cassette. So >we would be interested in other people's experience with this approach to >double KO in ES cells (and any tidbits on experience targeting the second >allele indepndantly).

For what it is worth, I sequenced Mortensen's pNTK construct, which I am pretty sure was the one he used in the MCB paper, and it was of the mutant neo variety. I am at home at present, but if you haven't seen it, then John Sedivy had a very interesting MCB a year or so back on titrating G418 to obtain high efficiency KO's, with some interesting observations that related to the homozygous KO method. Let me know if you need more info, and I'll dig it out when I am at work. I have an oligo from close to the polymorphic site in the neo gene, if you want to check other neo constructs by sequencing - you're welcome to some of it.

Cheers,

Martin

NNNN NN Martin A Kennedy (E-mail = [email protected]) ZZZZZZZ NN NN NN Cytogenetic and Molecular Oncology Unit ZZZ NN NN NN Christchurch School of Medicine ZZZ NN NNNN Christchurch, New Zealand ZZZZZZZ Phone (64-3)364-0880 Fax (64-3)364-0750

Subject: Knockout advice -Reply Message-ID:

Hi Sally--

We use Swiss Webster vasectomized males, bought from Taconic. We don't routinely test them, and have so far (knock on wood!) never had a leak. When I used to do the vasectomies myself, we tested them with two females each, let them plug both, then waited 3 weeks for hopefully no resultant pregnancies. We use vasectomized males twice a week to every other day. They can be used every night for a couple of weeks before needing some down time. Once a week is ridiculous, they don't need to keep their sperm count up after all! We use them for a year as well, or until they stop performing. We use Swiss Webster or CD-1 females for recipients--they are easy to estrus select, good mothers, and cheap. I wouldn't keep them on a high fat diet--in fact, we routinely cull out recipients that are too fat and discard them.

As for testing germline transmission, 100 pups to "prove" lack of transmission is the standard, but practicality usually dicatates compromise. We only breed males over 50% chimeric, females only if there are no male chimeras available. Three litters is a good number, if they're good-sized litters, I usually try for 25-30 pups.

We are using E14.1 cells from Cincinnati, but I've used CCE, D3, J1, R1, etc. Cell line is crucial; I've heard Genome Systems' line works fine.

We superovulate for blastocysts routinely, using C57Bl6 females. We generally do 10 at a time, with the standard amounts/timing of hormones, and a 6:30am/6:30pm light cycle. The one thing we find is crucial is to match body weight of the donors, 13-15g at time of PMS administration. We do three injection days per clone, averaging 30 blastocysts per 10 donors (with the occasional null day). I have gotten good results with number and % chimerism, as well as germline transmission with these blastocysts. We only use the males once per week, and usually only for 6-8 months, though a colleague says she uses them twice a week for a year with no trouble.

Good Luck-- Valerie Stewart Transgenic/Knockout Facility Cleveland Clinic Research Institute "[email protected]"

>>> Sally Wixson, VMD, MS - 8/13/96 1:16 PM >>> I direct the Lab Animal department of a pharmaceutical research company that has an active, but disorganized, knock out team. I have been here eight months and trying to get their animal work back on track. I would be grateful for your advice on several topics: .....what strain of vactomized males do you use and why? How do you assess their sterility (other than ordering then vasec. by the vendor)? How often do you use vasec. males? (my invesitagors use each male once/week or less!)

From crussell at ResGen.COM Mon Aug 19 17:03:24 1996 From: crussell at ResGen.COM (Chris Russell) Subject: employment opportunity Message-ID: <[email protected]>

Position Available Immediately! Lab Animal Technologist/Research Assistant

Skills Required: --Knowledgeable in theoretical embryology --Skilled in applied embryology and microsurgery techniques --Primarily laboratory work using the mouse as a model system --Molecular biology experience preferred

--Salary: $28-30K plus great benefits

--Excellent work environment in a growing biotech firm in Huntsville, AL

Please send resume and references to: Dr. Chris Russell Research Genetics, Inc. 2700 Memorial Parkway, SW Huntsville, AL 35801 1-800-711-2089 e-mail: [email protected] *********************************************************************** **** Christopher G. Russell, Ph.D. RESEARCH GENETICS, INC Resources for Research -_+ _+_+ ++-_ _+-_ _+_ _++ _+-_ -+ -+ -+ +- +- -+ + -+ -+ -+ + -+ -+ -+ -+ +- -+ -+ -+ -+ -+ -+ -+ -+ -++-+ -+_-+ -+_+ -++- -+_++ -+_+ 2700 Memorial Parkway SW Huntsville, AL 35801 Phone 1-800-533-4363, ext 2211 or Direct 1-800-711-2089 U.K. 0-800-89-1393 Fax 1-205-551-1021 or 1-205-536-9016 [email protected] *********************************************************************** ******

Subject: blastocyst culture -Reply Message-ID:

Karin--

We use 5 units PMS and HCG administered at 2pm and 12pm respectively. Our facility is on a 12 hour cycle, 6:30am-6:30pm. I've found that lower amounts of hormone work just as well, and sometimes better with C57s. Also I've found that using donors selected for body weight (13-15g at time of PMS injection) helps rather than using mice the same age and wildly different sizes. The number of blastocysts still varies day to day, but averages out over time. We inject 2-3 days per clone, and get plenty of chimeras. We flush blastocysts, and I wouldn't flush earlier than morulae--culture conditions can be tricky.

>>> Karin van Veen - 8/15/96 6:52 AM >>>

Hi, you all!

From kveen at nki.nl Tue Aug 20 10:17:30 1996 From: kveen at nki.nl (Karin van Veen)

Subject: blastocyst culture THANKS Message-ID:

Hello to everybody!

I want to thank everybody that reacted at my call for help concerning blastocyst-yield problems. Overall I can say that we are not doing things very wrong, most people use more or less the same techniques and time scedules. With some minor variations here and there. There where some tips I'm surely gonna try! I was glad to hear that almost everybody has or had the same problems as we have. Blastocyst yields from B6 mice varies a lot! It goes better and worse, without any clear reason. Last week I had a good week again, on three injection days, using 40 superovulated mice, I made 8 fosters, so almost 2 clones! I know, it is not fantastic, but I was happy. I'll let you know when I find THE answer to the problems. Bye, Karin. The Netherlands Cancer Institute Amsterdam

From tjf at uci.edu Tue Aug 20 17:28:50 1996 From: tjf at uci.edu (Tom Fielder)

Subject: Pricing Message-ID:

I am setting up a transgenic mouse core facility at UC-Irvine. We are trying to decide on our recharge rates, and I was hoping to get some feedback as to what other transgenic core facilities charge. We are planning to offer 2 levels of service. The basic package would be a fee per round of injection, with no guarantees as to outcome. The deluxe package would be a larger fee but would guarantee a certain number of transgenic pups (maybe 2?) (with perhaps a limit on the total number of tries). I'll try to extract as much info as I can from web pages, so if your prices are clearly posted (and readily accessible, e.g., through the Rat and Mouse Home Page, etc.), don't bother to respond. You can make it very brief, with just the price and what's guaranteed, and I will summarize the results for the list (minus names). I will post this same request on the Embryo Mail List and Compmed. Thanks!! Tom

From magree00 at pop.uky.edu Wed Aug 21 14:27:02 1996 From: magree00 at pop.uky.edu (Mike Green)

Subject: DNA quality/concentration Message-ID: <[email protected]>

When one of our researchers decides to produce transgenic mice, we provide a standard protocol for preparing the DNA for microinjection. In the past we've had a small number of labs who have produced generally high quality DNA for microinjection. Now that the number of facility users is increasing, we're implementing a pre-injection screening of the DNA to be injected. We plan to run a mini-gel to verify that there is a single band of the size and concentration claimed by the researcher.

Do other facilities do pre-injection screening? If so, what do you look for?

Would a spectrometer reading also be useful? Since most samples are of low volume and low concentration, I've been looking into the 7ul "ultra microvolume" cells for our GeneQuant. Does anyone have experience with these?

Mike Green University of Kentucky Transgenic Facility [email protected] 333 Combs phone: 606.257.2118 800 Rose St fax: 606.257.8940 Lexington, KY 40536 http://www.uky.edu/Transgenic/

Subject: DNA quality/concentration Message-ID:

I would certainly recommend gel checking for both band size and DNA concentration. Lately I noticed that such purified DNA when stored at 4oC for a long time (over 12 months) are no longer able to produce transgenic mice by pronuclei injection. This is true even for DNA samples that have been proven to generated transgenic founders. Has anyone had the same experience?

>When one of our researchers decides to produce transgenic mice, we provide a >standard protocol for preparing the DNA for microinjection. In the past >we've had a small number of labs who have produced generally high quality >DNA for microinjection. Now that the number of facility users is increasing, >we're implementing a pre-injection screening of the DNA to be injected. We >plan to run a mini-gel to verify that there is a single band of the size and >concentration claimed by the researcher. > >Do other facilities do pre-injection screening? If so, what do you look for? > >Would a spectrometer reading also be useful? Since most samples are of low >volume and low concentration, I've been looking into the 7ul "ultra >microvolume" cells for our GeneQuant. Does anyone have experience with these? > > >

Seong-Seng Tan

======

From pinkert at cmed.bhs.uab.edu Thu Aug 22 07:43:11 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert)

> Date: Thu, 22 Aug 1996 13:17:47 +1100 > To: [email protected] > From: [email protected] (Seong Seng Tan) > Subject: Re: DNA quality/concentration > Reply-to: [email protected]

> I would certainly recommend gel checking for both band size and DNA > concentration. Lately I noticed that such purified DNA when stored at 4oC > for a long time (over 12 months) are no longer able to produce transgenic > mice by pronuclei injection. This is true even for DNA samples that have > been proven to generated transgenic founders. Has anyone had the same > experience? No we haven't seen an absolute result like this. What does the gel checking show you after a year? Is there degradation, additional bands, or just a concentration difference? Although we too have had doubts about long-term storage, we routinely store aliquots at -20 and we have used an aliquot of a 'test' gene over a two year period successfully. C.A. Pinkert

From Allison_F._Treloar at ccmail.bms.com Thu Aug 22 14:09:07 1996 From: Allison_F._Treloar at ccmail.bms.com (Allison F. Treloar)

>>...Lately I noticed that such purified DNA when stored at 4oC for a long time (over 12 months) are no longer able to produce transgenic mice by pronuclei injection. This is true even for DNA samples that have been proven to generated transgenic founders. Has anyone had the same experience? ------

Absolutely! We routinely do a quick agarose check gel on stocks after 6 months to make sure they haven't degraded (noted as multiple bands or ladders). To keep our stocks fresh, we also keep an aliquot frozen just in case there is degredation in our working solution. (Don't freeze/thaw your stocks often, though!) Depending on your assay, you may still get positives, even with degraded samples, but they won't be expressors because the whole gene construct probably isn't intact.

In response to S-S Tan's questions: We have the investigator send us a gel picture of the construct band to show a single band is indeed present. We don't require a spec reading, but it's a nice addition. Have the investigator supply it! It sure is a lot easier to have them do it than to spend money buying yourself new equipment.

Allison Treloar Technical Supervisor - Transgenics Bristol-Myers Squibb [email protected]

"I try to take one day at a time, but sometimes several days attack me at once." From kelly at citi2.fr Fri Aug 23 01:12:18 1996 From: kelly at citi2.fr (U344)

Subject: 129SV Peculiarities... Message-ID:

Hello, we've got K.O. mice which are 129/bl6 hybrids, and are trying to get our phenotypes back onto 129. Problem is, we're having a hard time getting these animals to reproduce... very few, pregnancies, then very low litter yields, or many pup deaths. Does anyone have experience with 129SV mice who could let me know if these peculiarities are common with this strain, or what I could do to make them happier? Any help would be greatly appreciated.

cheers...

From crussell at ResGen.COM Thu Aug 22 15:23:32 1996 From: crussell at ResGen.COM (Chris Russell)

Subject: 129SV Peculiarities... Message-ID: <[email protected]>

Some info from another mouse list:

We have backcrossed a 129 ES cells derived knock-out mice into B6. After 5 generations we ran into problems of very low fertility. We used good B6 partners for all mating, thus the problem is not B6.

Decrease in fertility is common in intercrosses during the production of congenic lines but are not supposed to happen in backcrosses. During the 5th and 6th generations of our backcross, we only obtained 1- 2 successiful litters of 4-6 animals after a year of mating each offspring with a healthy B6 partner. Anyway we were able to survive the crisis, and after the 7th backcross generation the fertility problem disappeared.

Genetic heterogeniety effects in KO mice is getting more noticed. Thus it is wise to carefully plan one's mating strategies or one's choice of ES cells.

> Hello, we've got K.O. mice which are 129/bl6 hybrids, and are >trying to get our phenotypes back onto 129. Problem is, we're having a >hard time getting these animals to reproduce... very few, pregnancies, then >very low litter yields, or many pup deaths. Does anyone have experience >with 129SV mice who could let me know if these peculiarities are common >with this strain, or what I could do to make them happier? Any help would >be greatly appreciated. > > cheers... > >INSERM Unite 344 Endocrinologie Moleculaire >156 rue de Vaugirard, 75730 Paris cedex 15 France > >'Man... I don't EVEN have an opinion...' > > > > > > > *********************************************************************** **** Christopher G. Russell, Ph.D. RESEARCH GENETICS, INC Resources for Research -_+ _+_+ ++-_ _+-_ _+_ _++ _+-_ -+ -+ -+ +- +- -+ + -+ -+ -+ + -+ -+ -+ -+ +- -+ -+ -+ -+ -+ -+ -+ -+ -++-+ -+_-+ -+_+ -++- -+_++ -+_+ 2700 Memorial Parkway SW Huntsville, AL 35801 Phone 1-800-533-4363, ext 2211 or Direct 1-800-711-2089 U.K. 0-800-89-1393 Fax 1-205-551-1021 or 1-205-536-9016 [email protected] *********************************************************************** ****** From Allison_F._Treloar at ccmail.bms.com Thu Aug 22 17:04:43 1996 From: Allison_F._Treloar at ccmail.bms.com (Allison F. Treloar)

Subject: 129SV Peculiarities... Message-ID: <[email protected]>

>129/bl6 hybrids....very few, pregnancies, then very low litter yields, or many pup deaths .

129 females are not good mothers to start off with. All the problems you describe above are common to this strain. I don't know how to increase the pup yields (superovulation?), but you can combat the pup death issue by fostering off the pups to better ICR moms right away. We keep a few foster moms going for this purpose (about 2 foster litters drop/week). Couldn't hurt ;)

Allison Treloar Technical Supervisor-Transgenics Bristol-Myers Squibb Co.

Subject: Source for M2 medium Message-ID:

Help! Does anyone know of a source for M2 medium (for manipulation of mouse eggs) besides Sigma? They're back-ordered for 3-4 weeks. (Yes, I have the recipe, but not all the ingredients, nor the time to run quality control on home-made stuff). Tom

Thomas J. Fielder UC-Irvine University Lab Animal Resources Transgenic Mouse Facility 147 BSA Irvine, CA 92697-1310 e-mail: [email protected] phone: 714-824-8579 FAX: 714-824-2003 From browng at medicine.wustl.edu Fri Aug 23 12:58:44 1996 From: browng at medicine.wustl.edu (Gary Brown)

Subject: Source for M2 medium In-Reply-To: Message-ID:

I think that Gibco-BRL have M2, M16, BMOC (possibly more). Check out this source.

Enjoy,

On Thu, 22 Aug 1996, Tom Fielder wrote:

> Help! Does anyone know of a source for M2 medium (for manipulation of > mouse eggs) besides Sigma? They're back-ordered for 3-4 weeks. (Yes, I > have the recipe, but not all the ingredients, nor the time to run quality > control on home-made stuff). > Tom > > Thomas J. Fielder > UC-Irvine > University Lab Animal Resources > Transgenic Mouse Facility > 147 BSA > Irvine, CA 92697-1310 > e-mail: [email protected] > phone: 714-824-8579 > FAX: 714-824-2003 > > > > >

From Allison_F._Treloar at ccmail.bms.com Fri Aug 23 14:11:00 1996 From: Allison_F._Treloar at ccmail.bms.com (Allison F. Treloar)

Subject: Source for M2 medium Message-ID: <[email protected]> >Help! Does anyone know of a source for M2 medium (for manipulation of mouse eggs) besides Sigma?

I will be one of the hundreds of replies you'll receive who will recommend Specialty Media, Inc. 800.543.6029

They supply M2 (and other common media) in liquid or lyophilized powder (my favorite because of the longer storage time).

liquid (50ml) MR-015-D $16.50 powder (5x10ml) MR-015P-5F $40.00

Good luck.

Allison

From mbo at avery.med.virginia.edu Fri Aug 23 13:26:07 1996 From: mbo at avery.med.virginia.edu (M. B. Ober)

Subject: Source for M2 medium Message-ID:

I get my M2/M16 from Specialty Media, 1-800-543-6029. They have ready-to-thaw and rehydratable versions.

Subject: Source for M2 medium Message-ID: <[email protected]>

Gary, I had already checked the Gibco catalog and didn't see any of these (although I didn't call). However, 4 other people have already recommended Specialty Media, Inc. in New Jersey (800-543-6029). Fortunately, they have it in stock, although at more than twice the cost of Sigma. Tom

>I think that Gibco-BRL have M2, M16, BMOC (possibly more). Check out this >source. > >Enjoy, > >Gary Brown >E-mail: [email protected] or [email protected] >Web: http://ourworld.compuserve.com/homepages/TheBroons/ > "The Microinjection Workshop" > > >On Thu, 22 Aug 1996, Tom Fielder wrote: > >> Help! Does anyone know of a source for M2 medium (for manipulation of >> mouse eggs) besides Sigma? They're back-ordered for 3-4 weeks. (Yes, I >> have the recipe, but not all the ingredients, nor the time to run quality >> control on home-made stuff). >> Tom >> >> Thomas J. Fielder >> UC-Irvine >> University Lab Animal Resources >> Transgenic Mouse Facility >> 147 BSA >> Irvine, CA 92697-1310 >> e-mail: [email protected] >> phone: 714-824-8579 >> FAX: 714-824-2003 >> >> >> >> >> > > > > > Thomas J. Fielder UC-Irvine University Lab Animal Resources 147 BSA Irvine, CA 92697-1310 [email protected] From mbo at avery.med.virginia.edu Mon Aug 26 16:45:23 1996 From: mbo at avery.med.virginia.edu (M. B. Ober)

Subject: PMS Gonadotropin Message-ID:

From sp3i at avery.med.virginia.edu Mon Aug 26 16:58:41 1996 From: sp3i at avery.med.virginia.edu ([email protected])

Subject: PMS Gonadotropin Message-ID:

M >I've been using the 50 IU/vial lyophyllized PMSG. Is this overkill? When >Sigma had their 50IU/vial hCG on backorder I took a chance on ICN's hCG >(5000 IU for the price of one Sigma vial) and it seems to be working fine. >Can I get away with cheaper PMSG also?

It seems there's nothing to lose by trying the cheaper PMSG if you can get/find it.

Is it the case that 22 out of 28 hostesses were pregnant in the SM22 series? That's my read from the database. S

------Sonia Pearson-White, Ph.D. 804-982-0756 tel., 804-982-3993 fax. University of Virginia Medical Center, MR4 Room 3002, 300 Park Place Charlottesville, VA 22908

From PARLOW at AFP76.HUMC.EDU Mon Aug 26 17:30:50 1996 From: PARLOW at AFP76.HUMC.EDU (phone(310)222-3537 fax222-3432)

Subject: PMS GONADOTROPIN AVAILABLE Message-ID: <[email protected]>

REGARDING SUPPLY OF PMSG, TWO CONCERNS ARE

1) ACCURACY OF THE STANDARDIZATION 2) COST

PMSG IS CURRENTLY AVAILABLE FROM MY LABORATORY IN A RELIABLY STANDARDIZED FORMAT, IN ABUNDANT SUPPLY.

REQUESTS MAY BE SUBMITTED TO MY E-MAIL ADDRESS OR TO MY FAX LISTED ABOVE OR BY MAIL TO

DR. A. F. PARLOW HARBOR-UCLA MEDICAL CENTER PITUITARY HORMONES/ANTISERA CENTER 1000 WEST CARSON ST. TORRANCE, CA 90509

I WILL COME TO YOUR RESCUE.

======your inquiry appears below ======

From: IN%"[email protected]" 26-AUG-1996 08:47:25.05 To: IN%"[email protected]" CC: Subj: PMS Gonadotropin

From jparkert at magnus.acs.ohio-state.edu Mon Aug 26 19:55:58 1996 From: jparkert at magnus.acs.ohio-state.edu (Janice Parker-Thornburg)

I guarantee a set number of transgenics with each injection, and have found the DNA prep to be critical. Thus, rather than trust some of the researchers, I request a restriction digest along with a picture proving it is digested, and where the band of interest is circled. Then, I prep the DNA myself. I have had very good luck with this. No, I would not trust many researchers to prepare high quality DNA--especially if they have never done a microinjection before. As a molecular biologist myself, I know that one is often less than careful when prepping DNA for cloning and digests. However, as an injector, I also know how pure the DNA must be for this type of work. If you can, do it yourself. I can send you an easy prep that gives great DNA.

From h-marsha at nimr.mrc.ac.uk Tue Aug 27 09:42:39 1996 From: h-marsha at nimr.mrc.ac.uk (Heather)

Subject: PMS Gonadotropin Message-ID: >Sigma has just informed me that their Pregnant Mares' Serum Gonadotropin is >on backorder. Last time they did this it was almost 6 months before they >had it available. What is a good alternative source? > >I've been using the 50 IU/vial lyophyllized PMSG. Is this overkill? When >Sigma had their 50IU/vial hCG on backorder I took a chance on ICN's hCG >(5000 IU for the price of one Sigma vial) and it seems to be working fine. >Can I get away with cheaper PMSG also? > >Maggie Ober >Transgenic Mouse Core Facility >University of Virginia

Hello Maggie,

This may not help you much as it involves shipping costs, but if you're really stuck, then Intervet Labs in Cambridge, UK. may be able to help. They provide a box of 5 vials PMS (1000IU/vial) for ?17.40. This works out at about ?3.50 per 1000 IU, where Sigma charge ?33.70 for the same amount!! Intervet's hCG is also supplied at 5 vials per box, but at 1500IU per vial (?38 per box). I can give you their details if you want to take this further.

Heather Marshall Division of Developmental Neurobiology National Institute for Medical Research The Ridgeway Mill Hill London. NW7 1AA Tel:(0181-959-3666) [email protected]

From jparkert at magnus.acs.ohio-state.edu Thu Aug 29 20:58:53 1996 From: jparkert at magnus.acs.ohio-state.edu (Janice Parker-Thornburg)

Subject: B-cell specific expression Message-ID: <[email protected]>

This is just a quick query to see if I can get a response here prior to doing some library legwork. I am working with an investigator who wants to have his gene expressed in B-cells in transgenic mice. I was wondering if anyone out there either has, or knows of someone who has a plasmid containing the mouse IgG enhancer with an appropriate promoter and subsequent cloning sites that has worked well in transgenics. The one he has now has an IgG enhancer of unknown origin followed by the promoter for a housekeeping gene. I'm rather skeptical that it will work to the extent that he needs it. I appreciate any leads.

Thanks much.

From Dnxrh at aol.com Thu Aug 29 21:35:53 1996 From: Dnxrh at aol.com ([email protected])

Subject: B-cell specific expression Message-ID: <[email protected]>

You can do an on-line keyword search (without leaving your seat) of the Oakridge National Laboratories Transgenic and Targeted Mouse Database at: http://www.ornl.gov/TechResources/Trans/hmepg.html

One suggestion. Once you have identified potential promotors by doing a "B-Cell" keyword search, you might want to go back and use the promotor itself as a keyword for a follow up search. You never know what other tissues a particular promotor might also express in and that can be relevant.

Rick Huntress DNX Transgenics [email protected] 508-779-0189 (phone) 508-779-0190 (fax)

1996 September From DAMAK at whio.lincoln.ac.nz Mon Sep 2 14:32:54 1996 From: DAMAK at whio.lincoln.ac.nz (Damak, Sami) Date: Wed Aug 20 11:52:52 2003 Subject: Import of transgenic mice into the US Message-ID: <[email protected]>

I am looking at sending transgenic mice from New Zealand to the US. Does anyone know who I should approach to get customs and quarantine clearance? Thanks Sami Damak

From Dnxrh at aol.com Tue Sep 3 00:07:29 1996 From: Dnxrh at aol.com ([email protected]) Date: Wed Aug 20 11:52:52 2003 Subject: Import of transgenic mice into the US (DNA Too!) Message-ID: <[email protected]>

If permits are required (they may not be) it is the responsibility of the importer (your US recipient) to get the permits. Have them contact the USDA - APHIS to determine if permits are needed.

USDA - APHIS can be found at: http://www.aphis.usda.gov 301-734-7885 (phone) 301-734-8226

The guidelines seem to change but at the present time you should know the following:

Do the mice contain any livestock or avian genes ?

Do the mice contain and genes from any livestock or avian pathogens?

Are the mice infected with any known diseases which are transferable to humans, birds or livestock?

Do the mice contain any genes from human pathogens?

Have the mice been treated with any human blood products or any livestock derived products (ie. albumin).

If you can answer "no" to these questions then your recipient in the US will probably will not need a permit. Even if a permit is not required, the animals will still need to have paperwork documenting where they come from, what genes they contain and what their current health status is. If you do need a permit they can take 6 weeks to get.

I have had more experience shipping mice out of the US than in shipping them into the US. I suggest very strongly that you or your recipient in the US contact USDA-APHIS directly to confirm this information.

PS. to all you gene jockeys out there. These same rules apply to all recombinant DNA as well. We have had plasmids and DNA fragments held up by customs because the invoices did not state clearly all of the above.

Good Luck, Rick Huntress DNX Transgenics [email protected] +508-779-0189 (phone) +508-779-0190 (fax)

From WRIGHTA at gunet.georgetown.edu Tue Sep 3 12:07:11 1996 From: WRIGHTA at gunet.georgetown.edu (Ann Wright) Date: Wed Aug 20 11:52:52 2003 Subject: Import of transgenic mice into the US -Reply Message-ID:

Information regarding APHIS programs and import-export permits can be obtained on the home page http://www.aphis.usda.gov. The fax number is 301-734-8226, email [email protected] or voice mail 301-734-4412. The permit to import animals is needed to clear customs.

From carton at murray.fordham.edu Tue Sep 3 20:02:12 1996 From: carton at murray.fordham.edu (Jill Carton) Date: Wed Aug 20 11:52:52 2003 Subject: electroporators for ES cells Message-ID:

I am a graduate student at Fordham University working on generating a knockout mouse. The electroporation protocols I have found for the ES cells are all using the Biorad Gene Pulser. We have in our department a Baekon 2000 gene transfer system. I was wondering if anyone has ever used this apparatus for electroporation of ES cells and if you would share your protocol with me. This would save me a lot of time trying to work out the conditions.

Thanks in advance for the information, Jill Carton

Jill Carton Dept. Biological Sciences Fordham University Larkin Hall, rm 160 Bronx, NY 10458 718-817-3652 718-817-3645 (fax) [email protected]

From jklohse at facstaff.wisc.edu Fri Sep 6 18:57:44 1996 From: jklohse at facstaff.wisc.edu (Jan K. Lohse) Date: Wed Aug 20 11:52:52 2003 Subject: transgene control systems Message-ID:

I'm interested in finding out what sorts of transgenic mice people are currently creating with non-leaky, induceable promoter systems such as the ones that use the tet operon. Are these being used with good success? Satisfactory expression, turned on or off by a drug as designed? Is the lac operon being used for this sort of thing as well? Any descriptions of constructs, suggestions, and info about potential problems you're willing to share would be appreciated.

Also, how about similar info about recombination modification systems such as CRE-LOX and FLP?

Any other cool control systems out there?

Thanks very much. I think many of us will find the answers interesting.

From fmargoli at umabnet.ab.umd.edu Mon Sep 9 10:36:33 1996 From: fmargoli at umabnet.ab.umd.edu (Frank L. Margolis) Date: Wed Aug 20 11:52:52 2003 Subject: transgene control systems Message-ID:

>I'm interested in finding out what sorts of transgenic mice people are >currently creating with non-leaky, induceable promoter systems such as the >ones that use the tet operon. Are these being used with good success? >Satisfactory expression, turned on or off by a drug as designed? Is the >lac operon being used for this sort of thing as well? Any descriptions of >constructs, suggestions, and info about potential problems you're willing >to share would be appreciated. > >Also, how about similar info about recombination modification systems such >as CRE-LOX and FLP? > >Any other cool control systems out there? > >Thanks very much. I think many of us will find the answers interesting. > >Jan Similar questions arise with regard to the ecdysone regulated promoter recently reported in PNAS. Any info would be useful to all.

Frank Margolis

"There is something fascinating about science: one gets such wholesale returns of conjecture out of such a trifling investment of fact." Mark Twain

From Roger.Leemann at imr.psi.ch Mon Sep 9 20:37:54 1996 From: Roger.Leemann at imr.psi.ch (Roger C. Leemann) Date: Wed Aug 20 11:52:52 2003 Subject: visibility of cell nuclei in tw-cell mouse embryos Message-ID: <[email protected]>

Gary Brown came across a few questions I had sent to the rodent-research list and because he had to admit that his answers to most of them would be "I don't know" suggested I might give it a try on this list.

Sure, I'd like to, although it's only indirectly related to transgenic mice. Thanks Gary. I use 2-cell embryos of B6C3F1xB6C3F1 (Charles River, Germany) in a project where we want to study the effects of irradiation (e.g. alpha, protons) on the level of individual cells. We are able to detect single particle tracks with a track detector (basically a photographic emulsion) which is irradiated together with the cells or embryos with a vertical beam (normal to the plane of the detector). With a microscope we take pictures of both the cells or embryos and the developed detector and superimpose the two images. To tell which cell nuclei have been hit by a particle we must of course be able to see the boundary of the nucleus (we would like to get away without fluorescent staining). I have always had difficulties observing the nucleus in brightfield or phase contrast (we don't have Nomarski and Hoffman is no solution) in my B6C3F1 embryos. By chance I found out that in an other mouse strain (NMRI) the nucleus is clearly visible even with bright field illumination. Unfortunatly the NMRI embryos didn't develop beyond the 4-cell stage under my plain vanilla culture conditions which are OK for the B6C3F1.

My question therefore: Is anybody aware of a mouse strain, where the nuclei of life two-cell embryos are easily visible (unstained, no Nomarksi) and which is healthy enough, so that the embryos will develop in vitro to the blastocyst stage and hatch under standard culture conditions?

Any help is highly appreciated.

::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::: ::::: Work: Institute for Medical Radiobiology ++41 (56) 310-3792 [desk] CH-5232 Villigen PSI ++41 (1) 3856-561 [desk ZH] Switzerland ++41 (56) 310 3294 [fax] Home: Nordstrasse 26, CH-8006 Zuerich Switzerland ++41 (1) 361 0349 [home]

From pinkert at cmed.bhs.uab.edu Mon Sep 9 13:56:56 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert) Date: Wed Aug 20 11:52:52 2003 Subject: visibility of cell nuclei in tw-cell mouse embryos Message-ID: <19960909180142886.AAA77@cap1>

> Date: Mon, 9 Sep 1996 19:37:54 +0200 > To: [email protected] > From: [email protected] (Roger C. Leemann) > Subject: visibility of cell nuclei in tw-cell mouse embryos > Reply-to: [email protected]

> Hi > > Gary Brown came across a few questions I had sent to the rodent- research > list and because he had to admit that his answers to most of them would be > "I don't know" suggested I might give it a try on this list. > > Sure, I'd like to, although it's only indirectly related to transgenic > mice. Thanks Gary. > > I use 2-cell embryos of B6C3F1xB6C3F1 (Charles River, Germany) in a project > where we want to study the effects of irradiation (e.g. alpha, protons) on > the level of individual cells. We are able to detect single particle tracks > with a track detector (basically a photographic emulsion) which is > irradiated together with the cells or embryos with a vertical beam (normal > to the plane of the detector). > With a microscope we take pictures of both the cells or embryos and the > developed detector and superimpose the two images. To tell which cell > nuclei have been hit by a particle we must of course be able to see the > boundary of the nucleus (we would like to get away without fluorescent > staining). > I have always had difficulties observing the nucleus in brightfield or > phase contrast (we don't have Nomarski and Hoffman is no solution) in my > B6C3F1 embryos. By chance I found out that in an other mouse strain (NMRI) > the nucleus is clearly visible even with bright field illumination. > Unfortunatly the NMRI embryos didn't develop beyond the 4-cell stage under > my plain vanilla culture conditions which are OK for the B6C3F1. > > My question therefore: Is anybody aware of a mouse strain, where the nuclei > of life two-cell embryos are easily visible (unstained, no Nomarksi) and > which is healthy enough, so that the embryos will develop in vitro to the > blastocyst stage and hatch under standard culture conditions? > > Any help is highly appreciated. B6C3F1 ova are somewhat pigmented and a bit more difficult to work with than other strains. We've worked with a few different strains that have readily visible nuclei at the 2-cell stage. For a hybrid, B6SJL or B6D2 should work, also outbred Swiss (CD1, CF1, ICR, etc), as well as a host of other strains including inbreds, such as B6 and FVB.

From pinkert at cmed.bhs.uab.edu Mon Sep 9 14:01:18 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert) Date: Wed Aug 20 11:52:52 2003 Subject: (Fwd) Re: visibility of cell nuclei in tw-cell mouse embryos Message-ID: <19960909180604693.AAA95@cap1>

> My question therefore: Is anybody aware of a mouse strain, where the nuclei > of life two-cell embryos are easily visible (unstained, no Nomarksi) and > which is healthy enough, so that the embryos will develop in vitro to the > blastocyst stage and hatch under standard culture conditions? > > Any help is highly appreciated. B6C3F1 ova are somewhat pigmented and a bit more difficult to work with than other strains. We've worked with a few different strains that have readily visible nuclei at the 2-cell stage. For a hybrid, B6SJL or B6D2 should work, also outbred Swiss (CD1, CF1, ICR, etc.), as well as a host of other strains including inbreds, such as C57BL/6 and FVB - although the viability of the B6 ova would not be equivalent to the other strains. C.A. Pinkert

From tjf at uci.edu Mon Sep 9 12:40:31 1996 From: tjf at uci.edu (Tom Fielder) Date: Wed Aug 20 11:52:52 2003 Subject: fertilization of mouse eggs Message-ID:

I am using FVB and B6D2F1 mice to make transgenics. I just started superovulating the B6D2's last week, and I intended to mate them with B6D2 males, but since the males weren't quite old enough, I decided to mate them with some proven FVB studs. I've done this twice, and both times I got a very low percentage of fertilized eggs. Both times there were obvious plugs in 5/7 females, and the yield of eggs per oviduct varied quite a bit from mouse to mouse. The first time, 20% of the eggs were fertilized, and the second time only 10%, as judged by the presence of 2 pronuclei. Is there some reason why the B6D2F1 X FVB mating would produce this result? The males are not more than 5 months old and have produced very high percentages of fertilized eggs when mated with FVB females. All other factors are equal (same lot of gonadotropins, same room, same feed, same light cycle, same timing of injections). Any ideas or comments would be greatly appreciated. Tom From dbowtell at petermac.unimelb.edu.au Wed Sep 11 10:02:38 1996 From: dbowtell at petermac.unimelb.edu.au (David Bowtell) Date: Wed Aug 20 11:52:52 2003 Subject: Mating of chimeric mice Message-ID:

We have a dozen or so chimeric mice from a KO in W9.5 cells. There is a strong sex distortion and some of the mice are very strong. This line has been used very successfully in a collaborating laboratory, with a high frequency of germline transmission. The first few mice have been mated but have not plugged C57BL/6J female over a period of about 4 weeks (these chimeric males are now ca. 12wo). Clearly we have a way to go before we have a problem but does anyone have any comments on 'wringing' the most out of reluctant chimeras?

Thanks.

From pinkert at cmed.bhs.uab.edu Tue Sep 10 19:18:00 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert) Date: Wed Aug 20 11:52:52 2003 Subject: Mating of chimeric mice Message-ID: <19960910232244761.AAA62@cap1>

> have not plugged C57BL/6J female over a period of about 4 weeks (these > chimeric males are now ca. 12wo). Clearly we have a way to go before we > have a problem but does anyone have any comments on 'wringing' the most out > of reluctant chimeras? Replace the females with new mice (preferably in proestrus) or setting up for an IVF protocol (using survival surgery and removal of the vas deferens and epididymis from one side only - thereby maintaining the male for future breeding, and if there is a problem with the first attempt - you still have another chance at rescuing the line). The IVF scheme can also provide an index of relative fertility based on sperm concentration, morphology, and motility. Carl A. Pinkert

From TSAUNDER at hg-basic1mail.hg.med.umich.edu Wed Sep 11 09:34:42 1996 From: TSAUNDER at hg-basic1mail.hg.med.umich.edu (TSAUNDER) Date: Wed Aug 20 11:52:52 2003 Subject: Mating of chimeric mice Message-ID: <[email protected]>

David,

Our appraoch to chimeras is to give them them many mating opportunities. To wit: rotate 2 young C57BL/6 females (6-8 weeks old) through the male's cage every two weeks. This assumes that a male will have enough sperm for one fertile mating per week. Removed females are housed in pairs until it is determined whether they are gravid. By the time the third pair of females is introduced, you should have weaned pups from the first pair. Alternatively, you may wish to consider the IVF approach suggested by Carl Pinkert.

Date: Wed, 11 Sep 1996 09:02:38 +1000 To: [email protected] From: David Bowtell Subject: Mating of chimeric mice Reply-to: [email protected]

Thanks.

From jparkert at magnus.acs.ohio-state.edu Wed Sep 11 09:41:57 1996 From: jparkert at magnus.acs.ohio-state.edu (Janice Parker-Thornburg) Date: Wed Aug 20 11:52:52 2003 Subject: Mating of chimeric mice Message-ID: <[email protected]>

>We have a dozen or so chimeric mice from a KO in W9.5 cells. There is a >strong sex distortion and some of the mice are very strong. This line has >been used very successfully in a collaborating laboratory, with a high >frequency of germline transmission. The first few mice have been mated but >have not plugged C57BL/6J female over a period of about 4 weeks (these >chimeric males are now ca. 12wo). Clearly we have a way to go before we >have a problem but does anyone have any comments on 'wringing' the most out >of reluctant chimeras?

>David Bowtell

David:

When we have "reluctant" males, our favorite trick is to give them one or two superovulated females, reasoning that: 1) we are sure that the females are in heat, and 2) the "quality" of their ovulatory phase might be better (of course, this is pure conjecture. . .).

Good luck.

Jan Parker-Thornburg, Manager Transgenic Animal Facility The Ohio State University [email protected] From carton at murray.fordham.edu Wed Sep 11 11:28:42 1996 From: carton at murray.fordham.edu (Jill Carton) Date: Wed Aug 20 11:52:52 2003 Subject: knockout mouse vector Message-ID:

Hi! I was wondering if anyone knows of a convenient vector for sale which could be used to generate a knockout transgene. I was thinking of a vector which contained the HSV-TK gene, and a neomycin resistance gene surrounded by two unique multiple cloning sites for homologous sequence insertion. Thanks.

From mlm at titus.u-strasbg.fr Thu Sep 12 12:20:39 1996 From: mlm at titus.u-strasbg.fr (Marianne LEMEUR) Date: Wed Aug 20 11:52:52 2003 Subject: NOD mice embryos Message-ID:

Has anyone an idea how to improve yields of NOD embryos. Superovulation is impossible and natural matings seem to be productive only in a narrow window.

Is there an hybrid cross which works well? We have tried B6XNOD which is not better.

Marianne Le Meur Transgenic Animal Facility IGBMC-Strasbourg, France From pinkert at cmed.bhs.uab.edu Thu Sep 12 09:59:26 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert) Date: Wed Aug 20 11:52:52 2003 Subject: NOD zygote yields Message-ID: <19960912140427737.AAA136@cap1>

> Has anyone an idea how to improve yields of NOD embryos. Superovulation is > impossible and natural matings seem to be productive only in a narrow > window. Marianne: As per Jackson Labs, superovulation actually works better in the NOD's with older females (evidently 8-12 weeks is better than 3-4 weeks). In our first attempt this year (with approx. 8 week old females), 3 of 23 females were plugged and yielded 60 injectable eggs, of which 29 survived, and resulting in 1 stillborn and 2 resorbing fetuses. We held mice for 3 more weeks, and in our second attempt, 7 of 28 females (including some a second time) were plugged, yielding 145 injectable eggs, of which 120 survived. There were 19 liveborn pups, 18 weaned, and 4 founder transgenics. (Last year 163 eggs were injected on 2 separate days, 132 survived and were transferred, yielding 7/29 transgenics, but we went through a goodly number of mice.) AI or IVF might be more beneficial, but the time to work out the additional conditions/scheduling combined with our poor IVF yields and variable timing with some BALB/c congenics and F1's dissuaded us from going ahead in that direction. Good luck. Carl

From TSAUNDER at hg-basic1mail.hg.med.umich.edu Fri Sep 13 08:31:13 1996 From: TSAUNDER at hg-basic1mail.hg.med.umich.edu (TSAUNDER) Date: Wed Aug 20 11:52:52 2003 Subject: Forwarded: Ovulation Model Message-ID: <[email protected]>

This message posted on behalf of Jennifer Bowen, Please direct replies to [email protected] -Thanks, Thom Saunders

I am interested in using a mouse model consisting of immature mice stimulated to ovulate a normal or near-normal number of follicles (ie not superovulated) and then mated to vasectomized males to induce pseudopregnancy. I am designing this model for future use in a mutant which is on a C57BL/6J X C3HB/FeJ background so will probably use C57BL/6 animals. I would appreciate any information on what sort of hormone dose/age combination will result in a normal number of ovulations in immature females of this or similar strains.

Jennifer Bowen ([email protected]) From mlm at titus.u-strasbg.fr Fri Sep 13 17:34:49 1996 From: mlm at titus.u-strasbg.fr (Marianne LEMEUR) Date: Wed Aug 20 11:52:52 2003 Subject: sending frozen embryos Message-ID:

I would like to send frozen embryos all over the world instead of sending living mice, which is very time consuming to get all the formalities together.

What are the conditions to do that, and were is it possible to buy the container. In France, our transportation company told us that the sender is personnally responsible in case of an accident with liquid nitrogen during the flight!

Marianne Le Meur IGBMC Strasbourg-France

From ucympek at ucl.ac.uk Fri Sep 13 17:47:32 1996 From: ucympek at ucl.ac.uk (Peter Koder) Date: Wed Aug 20 11:52:52 2003 Subject: sending frozen embryos Message-ID:

Hello Marianne

For a UK> recipient an Import licence is required from the Ministry of Agriculture tel (0)181 330 8178 under the Importation of Embryos, Ova and Semen order. Also the permission of a Home Office Inspector under the UK Animals(Scientific Procedures) Act 1986

The importing institute Genetically Modified Organism (GMO) safety committee has also to be informed inorder to accept them under the Environmental Protection Act (Contained Use) Regulations.

I hope this helps, Peter

I would like to send frozen embryos all over the world instead of sending >living mice, which is very time consuming to get all the formalities >together. > >What are the conditions to do that, and were is it possible to buy the >container. >In France, our transportation company told us that the sender is >personnally responsible in case of an accident with liquid nitrogen during >the flight! > >Marianne Le Meur >IGBMC >Strasbourg-France > > > > > >

Peter C Koder BVMS MSc MRCVS UCL Biological Services E-MAIL: [email protected] University College London TEL:+44(0)171 391 1309 Malet Place FAX:+44(0)171 380 7837 LONDON, WC1E 6BT UK

From pinkert at cmed.bhs.uab.edu Fri Sep 13 11:55:51 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert) Date: Wed Aug 20 11:52:52 2003 Subject: sending frozen embryos Message-ID: <19960913160057705.AAA159@cap1>

> Date: Fri, 13 Sep 1996 16:34:49 +0000 > To: [email protected] > From: [email protected] (Marianne LEMEUR) > Subject: sending frozen embryos > Reply-to: [email protected]

> I would like to send frozen embryos all over the world instead of sending > living mice, which is very time consuming to get all the formalities > together. > > What are the conditions to do that, and were is it possible to buy the > container. > In France, our transportation company told us that the sender is > personnally responsible in case of an accident with liquid nitrogen during > the flight! You may want to check with Dr. Glenn Monastersky regarding shipment of ova and related considerations. He was using the liquid nitrogen shipping containers that contained an absorbant material - it maintained temperature, yet there was no LN2 sloshing around at all. (what they used I believe was similar to the Arctic Express from Thermolyne (model CY50915) that keeps temperature for 11 days) . Glenn can be contacted at [email protected]. Carl

From andreja at ariel.ucs.unimelb.edu.au Sat Sep 14 11:09:28 1996 From: andreja at ariel.ucs.unimelb.edu.au (Andreja Pirkmaier) Date: Wed Aug 20 11:52:52 2003 Subject: Caesarean sections Message-ID: <[email protected]>

Has anyone any experience with injecting mice with hormones( Day, Dose) few Days prior doing caesarean section to avoid any early delivery. I know that the gestation period for the transgenic line I am interested in is 19 Days. I planed to do sections on Day 18 p. c..

Thanks Andreja Pirkmaier Peter MacCallum Cancer Institute [email protected]

From tonyjames at cuhk.edu.hk Sat Sep 14 10:43:18 1996 From: tonyjames at cuhk.edu.hk (A E James) Date: Wed Aug 20 11:52:52 2003 Subject: Caesarean sections Message-ID: <[email protected]>

> Has anyone any experience with injecting mice with hormones( Day, >Dose) few Days prior doing caesarean section to avoid any early delivery. > I know that the gestation period for the transgenic line I am >interested in is 19 Days. I planed to do sections on Day 18 p. c.. > >Thanks Andreja Pirkmaier >Peter MacCallum Cancer Institute >[email protected] > > > >Try Catherine O'Brien at WEHI or her senior technician at WEHI Kew...her name is Therese Johns. Any way my protocol is progesterone for three days before the expected birth. In pregnant rats the figure quoted is 20 mg per day for a heavy preg. rat. Other figures I have seen for mice are 1.5 to 2.5 mg per day per mouse for the three days prior to csaerian delivery. Hope this helps Tony James Tony James BVSc MACVSc Director Laboratory Animal Unit Chinese University of Hong Kong Shatin, New Territories, Hong Kong ph: 852 2609 6862 fx: 852 2603 5723 email: [email protected] ...... "nothing great was achieved without enthusiasm." R. W. Emerson

From pinkert at cmed.bhs.uab.edu Sat Sep 14 09:48:18 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert) Date: Wed Aug 20 11:52:52 2003 Subject: Caesarean sections Message-ID: <19960914135340470.AAA136@cap1>

> Has anyone any experience with injecting mice with hormones( Day, > Dose) few Days prior doing caesarean section to avoid any early delivery. > I know that the gestation period for the transgenic line I am > interested in is 19 Days. I planed to do sections on Day 18 p. c.. If the line uniformly delivers on d19 and the pups are robust, then C-sections on d18 should likely work out well without the additional hormone therapy.

From mlm at titus.u-strasbg.fr Tue Sep 17 16:19:26 1996 From: mlm at titus.u-strasbg.fr (Marianne LEMEUR) Date: Wed Aug 20 11:52:52 2003 Subject: zygote yields Message-ID:

Thanks to those who gave the solution for NOD mice zygote yield! More generally it seems that C57Bl6 is the only inbred strain which can be superovulated in the same way as an F1 hybrid. Other inbred strains like FVB/N or outbred strains like CD1 do not give better yields than can be achieved with a natural mating.Is that correct? Marianne Le Meur

From jparkert at magnus.acs.ohio-state.edu Tue Sep 17 11:45:24 1996 From: jparkert at magnus.acs.ohio-state.edu (Janice Parker-Thornburg) Date: Wed Aug 20 11:52:52 2003 Subject: zygote yields Message-ID: <[email protected]>

>Thanks to those who gave the solution for NOD mice zygote yield! >More generally it seems that C57Bl6 is the only inbred strain which can be >superovulated in the same way as an F1 hybrid. >Other inbred strains like FVB/N or outbred strains like CD1 do not give >better yields than can be achieved with a natural mating.Is that correct? >Marianne Le Meur

Marianne:

This is not correct. I have superovulated FVB/N for microinjection, as well as for general harvesting of mouse embryos. If done in the correct time frame, one can get 8-10 more eggs (embryos) per female than with natural matings. I also know that people using CD1's for embryo harvesting will superovulate them, presumably because they get a better yield. If you find that your yield does not improve with superovulation, it is likely that your mouse-age or timespan for the hormone injections needs to be optimized.

From landel at medsci.udel.edu Tue Sep 17 12:37:11 1996 From: landel at medsci.udel.edu (Carlisle Landel) Date: Wed Aug 20 11:52:53 2003 Subject: zygote yields Message-ID: <[email protected]>

Marianne,

No, this isn't correct! I superovulate FVB/N routinely, and I'm pretty sure that colleagues of mine in the past have done it to CD1's, too, though I vaguely recall there was some trick with dosage or timing or something (but I may be wrong--this was many years ago).

Check your hormones for proper concentration, and check that the light controls in your facility are properly set, and make sure that you are injecting at or very near the middle of your light cycle.

Regards, Carlisle Landel Dept. of Clinical Science A.I. duPont Institute PO Box 269 Wilmington DE 19899 (302) 651-6873 [email protected] http://mdblmac.medsci.udel.edu/WEB/MDBL/Carlisle.html

From pinkert at cmed.bhs.uab.edu Tue Sep 17 10:23:08 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert) Date: Wed Aug 20 11:52:53 2003 Subject: zygote yields Message-ID: <19960917142758010.AAB158@cap1>

> More generally it seems that C57Bl6 is the only inbred strain which can be > superovulated in the same way as an F1 hybrid. I'm not sure what is meant here, but other inbred strains can be utilized, using the same general scheme and sometimes jogging the schedule a bit.

> Other inbred strains like FVB/N or outbred strains like CD1 do not give > better yields than can be achieved with a natural mating.Is that correct? It's a matter of degree, but I disagree with the last statement (unless you mean that hybrids provide better net yields than either inbreds and outbreds, which is true). In some labs, natural matings facilitate a specific routine. However, there are significant economies-of-scale when superovulation is employed (in relation to zygote yields and needed labor/space/etc.) when using the strains that you have mentioned.

From browng at medicine.wustl.edu Tue Sep 17 14:03:45 1996 From: browng at medicine.wustl.edu (Gary Brown) Date: Wed Aug 20 11:52:53 2003 Subject: zygote yields In-Reply-To: Message-ID:

On Tue, 17 Sep 1996, Marianne LEMEUR wrote:

> Thanks to those who gave the solution for NOD mice zygote yield! > More generally it seems that C57Bl6 is the only inbred strain which can be > superovulated in the same way as an F1 hybrid. > Other inbred strains like FVB/N or outbred strains like CD1 do not give > better yields than can be achieved with a natural mating.Is that correct? > Marianne Le Meur > > > > Not so. I used FVB/N strain mice for 3 1/2 years and superovulated with (in my opinion) a high degree of success. Since it's a larger mouse than B6C3F1 hybrids, we used 7.5IU hormones rather than 5IU, and a 47 hr gap between PMSG and HCG. Typical yields were 25-30 good (non fragmented) eggs per mouse. Can't say I've had experience with other strains other than C57Bl/6 and B6C3F1 hybrids. Actually, I lie. 129SvJ mice are dismal for superovulation - if anyone can give me a way to do these reproducibly I'll get you a beer (or three)!

From carboni at valoritech.com Tue Sep 17 16:25:09 1996 From: carboni at valoritech.com (Nick Carboni) Date: Wed Aug 20 11:52:53 2003 Subject: Wood's coculture technique Message-ID: <[email protected]>

Hi everybody,

Short message to know if one of you is currently working or has tried to work on wood's coculture technique.

I have spoke with some people, and it appears that it is rather difficult to acheive.

Has anyone any experience to share with me ??

Thanks in advance,

Nick ------Nick Carboni, VP Valoritech 1253, McGill College Av., room 620 Montreal, Quebec, Canada H3B 2Y5 Tel.: (514) 866-8271 / Fax: (514) 866-8272 E-Mail: [email protected] From browng at medicine.wustl.edu Wed Sep 18 08:26:12 1996 From: browng at medicine.wustl.edu (Gary Brown) Date: Wed Aug 20 11:52:53 2003 Subject: Wood's coculture technique In-Reply-To: <[email protected]> Message-ID:

On Tue, 17 Sep 1996, Nick Carboni wrote:

> Hi everybody, > > > Short message to know if one of you is currently working or has tried to > work on wood's coculture technique. > > I have spoke with some people, and it appears that it is rather difficult to > acheive. > > Has anyone any experience to share with me ?? > > Thanks in advance, > > Nick > Hi !

I tried Wood's co-culture technique around 3 years ago, but only had 1 recorded birth in 16 implanted mothers, and this pup was malformed. We were unable to determine the efficacy of the technique as the ES cell line that we used was obtained from another laboratory, and both their technique and the cell line itself were called into question. However, here's what I can tell you..

1. We found that the ES cells did not adhere to the de - ZP'd morulae / octoploid / tetraploid embryos as well as we thought the paper indicated.

2. Our understanding is that the technique's success is very ES cell dependent.

3. Our understanding is that the live birth rate is very low, making this a system better suited to in utero developmental studies.

Since this time, I have tried to find the time to explore non-injection methods of chimera production (with little success on the time front!). I have only managed to complete a pilot study of Nagy / Rossant's "sandwich" method where I was successful in fusing 2 morulae together in culture overnight in custom made dimples. This effort would have been more instructive if I had had access to some ES cells to "sandwich", but the fusions had a >90% success rate. For the reference on this methodology, do a Medline search on the authors above or check out the E-newsletter archive on my homepage for August / September.

Hope this is instructive! :-)

From ldeg at midway.uchicago.edu Wed Sep 18 21:28:26 1996 From: ldeg at midway.uchicago.edu (Linda Degenstein) Date: Wed Aug 20 11:52:53 2003 Subject: ES Cells from Balb C mice Message-ID: <[email protected]>

We are looking for ES Cells (germline proven) that are made from Balb C strain mice. Does anyone know if these are available?

Linda Degenstein Director UCCRC Transgenic/ES Cell Facility The University of Chicago [email protected]

From khelmuth at facstaff.wisc.edu Tue Sep 24 15:28:52 1996 From: khelmuth at facstaff.wisc.edu ([email protected]) Date: Wed Aug 20 11:52:53 2003 Subject: noise problems Message-ID:

Hi everyone:

I am looking for documentation that excess noise can affect the breeding and production of embryos from mice. Does anyone know of published work that looked at environmental affects on a mouse colony? Any citations would be appreciated.

Kathy Krentz Helmuth Transgenic Facility 425 Henry Mall Room 2210 Biotechnology Center Madison, WI 53706 (608) 265-2801 email:[email protected]

From mlp at informatics.jax.org Tue Sep 24 17:20:46 1996 From: mlp at informatics.jax.org (Moyha Lennon-Pierce) Date: Wed Aug 20 11:52:53 2003 Subject: Noise/reproduction Message-ID:

The following references might be useful.

I searched the Strain Characteristics Catalog in development at The Jackson Lab and did a quick search in NLM. Not much at all in the literature, but the following references may be useful.

Nawrot PS et al. Embryotoxicity of various noise stimuli in the mouse. Teratology 1980; 22(3):279-89. CF-1

Zakem HB, Alliston CW. The effects of noise level and elevated ambient temperatures upon selected reproductive traits in female Swiss-Webster mice. Lab Anim Sci 1974; 24:469-75. Strain SWR

Moyha Lennon-Pierce MGD Informatics [email protected]

>Hi everyone: > >I am looking for documentation that excess noise can affect the breeding >and production of embryos from mice. Does anyone know of published work >that looked at environmental affects on a mouse colony? Any citations >would be appreciated. > >Kathy > >Kathy Krentz Helmuth >Transgenic Facility >425 Henry Mall >Room 2210 Biotechnology Center >Madison, WI 53706 >(608) 265-2801 >email:[email protected] From landel at medsci.udel.edu Tue Sep 24 18:13:36 1996 From: landel at medsci.udel.edu (Carlisle Landel) Date: Wed Aug 20 11:52:53 2003 Subject: Noise/reproduction Message-ID: <[email protected]>

Kathy,

I saw a poster at a meeting a couple of weeks ago where they were using one of those ultrasonic mouse repellers to increase the percentage of embryo loss in CBAxDBA/2 model. If you want more info, I'll see if I can point you towards the authors.

Regards,

Carlisle Landel Dept. of Clinical Science duPont Hospital for Children PO Box 269 Wilmington DE 19899 (302) 651-6873 [email protected] http://mdblmac.medsci.udel.edu/WEB/MDBL/Carlisle.html

From Roger.Leemann at imr.psi.ch Wed Sep 25 17:40:47 1996 From: Roger.Leemann at imr.psi.ch (Roger C. Leemann) Date: Wed Aug 20 11:52:53 2003 Subject: noise problems Message-ID: <[email protected]>

Kathy Krentz Helmuth ([email protected]) wrote:

>I am looking for documentation that excess noise can affect the breeding >and production of embryos from mice.

Sorry, no literature, but I can tell you how we do it. In our animal housing facility there is always a radio playing at a low volume during the daylight cycle (maybe this can not be referred to as "excessive" noise.) I don't know whether or how this affects the mice, positive or negative. With non-superovulated B6C3F1 mice I can expect 15% to 25% plugs and 7 to 12 two-cell embryos/mouse. How does this compare to other labs?

Roger ::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::: ::::: Work: Institute for Medical Radiobiology ++41 (56) 310-3792 [desk] CH-5232 Villigen PSI ++41 (1) 3856-561 [desk ZH] Switzerland ++41 (56) 310 3294 [fax] Home: Nordstrasse 26, CH-8006 Zuerich Switzerland ++41 (1) 361 0349 [home]

From DAMAK at whio.lincoln.ac.nz Mon Sep 2 02:32:54 1996 From: DAMAK at whio.lincoln.ac.nz (Damak, Sami)

Subject: Import of transgenic mice into the US Message-ID: <[email protected]>

I am looking at sending transgenic mice from New Zealand to the US. Does anyone know who I should approach to get customs and quarantine clearance? Thanks Sami Damak

From Dnxrh at aol.com Tue Sep 3 04:07:29 1996 From: Dnxrh at aol.com ([email protected])

Subject: Import of transgenic mice into the US (DNA Too!) Message-ID: <[email protected]>

If permits are required (they may not be) it is the responsibility of the importer (your US recipient) to get the permits. Have them contact the USDA - APHIS to determine if permits are needed.

USDA - APHIS can be found at: http://www.aphis.usda.gov 301-734-7885 (phone) 301-734-8226

The guidelines seem to change but at the present time you should know the following:

Do the mice contain any livestock or avian genes ?

Do the mice contain and genes from any livestock or avian pathogens?

Are the mice infected with any known diseases which are transferable to humans, birds or livestock?

Do the mice contain any genes from human pathogens? Have the mice been treated with any human blood products or any livestock derived products (ie. albumin).

If you can answer "no" to these questions then your recipient in the US will probably will not need a permit. Even if a permit is not required, the animals will still need to have paperwork documenting where they come from, what genes they contain and what their current health status is.

If you do need a permit they can take 6 weeks to get.

I have had more experience shipping mice out of the US than in shipping them into the US. I suggest very strongly that you or your recipient in the US contact USDA-APHIS directly to confirm this information.

PS. to all you gene jockeys out there. These same rules apply to all recombinant DNA as well. We have had plasmids and DNA fragments held up by customs because the invoices did not state clearly all of the above.

Good Luck, Rick Huntress DNX Transgenics [email protected] +508-779-0189 (phone) +508-779-0190 (fax)

From WRIGHTA at gunet.georgetown.edu Tue Sep 3 17:07:11 1996 From: WRIGHTA at gunet.georgetown.edu (Ann Wright)

Subject: Import of transgenic mice into the US -Reply Message-ID:

Information regarding APHIS programs and import-export permits can be obtained on the home page http://www.aphis.usda.gov. The fax number is 301-734-8226, email [email protected] or voice mail 301-734-4412. The permit to import animals is needed to clear customs.

From carton at murray.fordham.edu Wed Sep 4 00:02:12 1996 From: carton at murray.fordham.edu (Jill Carton)

Subject: electroporators for ES cells Message-ID: I am a graduate student at Fordham University working on generating a knockout mouse. The electroporation protocols I have found for the ES cells are all using the Biorad Gene Pulser. We have in our department a Baekon 2000 gene transfer system. I was wondering if anyone has ever used this apparatus for electroporation of ES cells and if you would share your protocol with me. This would save me a lot of time trying to work out the conditions.

Thanks in advance for the information, Jill Carton

From jklohse at facstaff.wisc.edu Fri Sep 6 23:57:44 1996 From: jklohse at facstaff.wisc.edu (Jan K. Lohse)

Subject: transgene control systems Message-ID:

I'm interested in finding out what sorts of transgenic mice people are currently creating with non-leaky, induceable promoter systems such as the ones that use the tet operon. Are these being used with good success? Satisfactory expression, turned on or off by a drug as designed? Is the lac operon being used for this sort of thing as well? Any descriptions of constructs, suggestions, and info about potential problems you're willing to share would be appreciated.

Also, how about similar info about recombination modification systems such as CRE-LOX and FLP?

Any other cool control systems out there?

Thanks very much. I think many of us will find the answers interesting.

Jan From fmargoli at umabnet.ab.umd.edu Mon Sep 9 15:36:33 1996 From: fmargoli at umabnet.ab.umd.edu (Frank L. Margolis)

Subject: transgene control systems Message-ID:

>I'm interested in finding out what sorts of transgenic mice people are >currently creating with non-leaky, induceable promoter systems such as the >ones that use the tet operon. Are these being used with good success? >Satisfactory expression, turned on or off by a drug as designed? Is the >lac operon being used for this sort of thing as well? Any descriptions of >constructs, suggestions, and info about potential problems you're willing >to share would be appreciated. > >Also, how about similar info about recombination modification systems such >as CRE-LOX and FLP? > >Any other cool control systems out there? > >Thanks very much. I think many of us will find the answers interesting. > >Jan Similar questions arise with regard to the ecdysone regulated promoter recently reported in PNAS. Any info would be useful to all.

Frank Margolis

"There is something fascinating about science: one gets such wholesale returns of conjecture out of such a trifling investment of fact." Mark Twain

From Roger.Leemann at imr.psi.ch Mon Sep 9 18:37:54 1996 From: Roger.Leemann at imr.psi.ch (Roger C. Leemann) Subject: visibility of cell nuclei in tw-cell mouse embryos Message-ID: <[email protected]>

Gary Brown came across a few questions I had sent to the rodent-research list and because he had to admit that his answers to most of them would be "I don't know" suggested I might give it a try on this list.

Sure, I'd like to, although it's only indirectly related to transgenic mice. Thanks Gary.

I use 2-cell embryos of B6C3F1xB6C3F1 (Charles River, Germany) in a project where we want to study the effects of irradiation (e.g. alpha, protons) on the level of individual cells. We are able to detect single particle tracks with a track detector (basically a photographic emulsion) which is irradiated together with the cells or embryos with a vertical beam (normal to the plane of the detector). With a microscope we take pictures of both the cells or embryos and the developed detector and superimpose the two images. To tell which cell nuclei have been hit by a particle we must of course be able to see the boundary of the nucleus (we would like to get away without fluorescent staining). I have always had difficulties observing the nucleus in brightfield or phase contrast (we don't have Nomarski and Hoffman is no solution) in my B6C3F1 embryos. By chance I found out that in an other mouse strain (NMRI) the nucleus is clearly visible even with bright field illumination. Unfortunatly the NMRI embryos didn't develop beyond the 4-cell stage under my plain vanilla culture conditions which are OK for the B6C3F1.

My question therefore: Is anybody aware of a mouse strain, where the nuclei of life two-cell embryos are easily visible (unstained, no Nomarksi) and which is healthy enough, so that the embryos will develop in vitro to the blastocyst stage and hatch under standard culture conditions?

Any help is highly appreciated.

::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::: ::::: Work: Institute for Medical Radiobiology ++41 (56) 310-3792 [desk] CH-5232 Villigen PSI ++41 (1) 3856-561 [desk ZH] Switzerland ++41 (56) 310 3294 [fax] Home: Nordstrasse 26, CH-8006 Zuerich Switzerland ++41 (1) 361 0349 [home] From pinkert at cmed.bhs.uab.edu Mon Sep 9 13:56:56 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert)

Subject: visibility of cell nuclei in tw-cell mouse embryos Message-ID: <19960909180142886.AAA77@cap1>

> Date: Mon, 9 Sep 1996 19:37:54 +0200 > To: [email protected] > From: [email protected] (Roger C. Leemann) > Subject: visibility of cell nuclei in tw-cell mouse embryos > Reply-to: [email protected]

> Hi > > Gary Brown came across a few questions I had sent to the rodent- research > list and because he had to admit that his answers to most of them would be > "I don't know" suggested I might give it a try on this list. > > Sure, I'd like to, although it's only indirectly related to transgenic > mice. Thanks Gary. > > I use 2-cell embryos of B6C3F1xB6C3F1 (Charles River, Germany) in a project > where we want to study the effects of irradiation (e.g. alpha, protons) on > the level of individual cells. We are able to detect single particle tracks > with a track detector (basically a photographic emulsion) which is > irradiated together with the cells or embryos with a vertical beam (normal > to the plane of the detector). > With a microscope we take pictures of both the cells or embryos and the > developed detector and superimpose the two images. To tell which cell > nuclei have been hit by a particle we must of course be able to see the > boundary of the nucleus (we would like to get away without fluorescent > staining). > I have always had difficulties observing the nucleus in brightfield or > phase contrast (we don't have Nomarski and Hoffman is no solution) in my > B6C3F1 embryos. By chance I found out that in an other mouse strain (NMRI) > the nucleus is clearly visible even with bright field illumination. > Unfortunatly the NMRI embryos didn't develop beyond the 4-cell stage under > my plain vanilla culture conditions which are OK for the B6C3F1. > > My question therefore: Is anybody aware of a mouse strain, where the nuclei > of life two-cell embryos are easily visible (unstained, no Nomarksi) and > which is healthy enough, so that the embryos will develop in vitro to the > blastocyst stage and hatch under standard culture conditions? > > Any help is highly appreciated. B6C3F1 ova are somewhat pigmented and a bit more difficult to work with than other strains. We've worked with a few different strains that have readily visible nuclei at the 2-cell stage. For a hybrid, B6SJL or B6D2 should work, also outbred Swiss (CD1, CF1, ICR, etc), as well as a host of other strains including inbreds, such as B6 and FVB.

From pinkert at cmed.bhs.uab.edu Mon Sep 9 14:01:18 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert)

Subject: (Fwd) Re: visibility of cell nuclei in tw-cell mouse embryos Message-ID: <19960909180604693.AAA95@cap1>

> My question therefore: Is anybody aware of a mouse strain, where the nuclei > of life two-cell embryos are easily visible (unstained, no Nomarksi) and > which is healthy enough, so that the embryos will develop in vitro to the > blastocyst stage and hatch under standard culture conditions? > > Any help is highly appreciated. B6C3F1 ova are somewhat pigmented and a bit more difficult to work with than other strains. We've worked with a few different strains that have readily visible nuclei at the 2-cell stage. For a hybrid, B6SJL or B6D2 should work, also outbred Swiss (CD1, CF1, ICR, etc.), as well as a host of other strains including inbreds, such as C57BL/6 and FVB - although the viability of the B6 ova would not be equivalent to the other strains. C.A. Pinkert

From tjf at uci.edu Mon Sep 9 19:40:31 1996 From: tjf at uci.edu (Tom Fielder)

Subject: fertilization of mouse eggs Message-ID:

I am using FVB and B6D2F1 mice to make transgenics. I just started superovulating the B6D2's last week, and I intended to mate them with B6D2 males, but since the males weren't quite old enough, I decided to mate them with some proven FVB studs. I've done this twice, and both times I got a very low percentage of fertilized eggs. Both times there were obvious plugs in 5/7 females, and the yield of eggs per oviduct varied quite a bit from mouse to mouse. The first time, 20% of the eggs were fertilized, and the second time only 10%, as judged by the presence of 2 pronuclei. Is there some reason why the B6D2F1 X FVB mating would produce this result? The males are not more than 5 months old and have produced very high percentages of fertilized eggs when mated with FVB females. All other factors are equal (same lot of gonadotropins, same room, same feed, same light cycle, same timing of injections). Any ideas or comments would be greatly appreciated. Tom

From dbowtell at petermac.unimelb.edu.au Wed Sep 11 00:02:38 1996 From: dbowtell at petermac.unimelb.edu.au (David Bowtell)

Subject: Mating of chimeric mice Message-ID:

Thanks.

From pinkert at cmed.bhs.uab.edu Tue Sep 10 19:18:00 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert) Subject: Mating of chimeric mice Message-ID: <19960910232244761.AAA62@cap1>

> have not plugged C57BL/6J female over a period of about 4 weeks (these > chimeric males are now ca. 12wo). Clearly we have a way to go before we > have a problem but does anyone have any comments on 'wringing' the most out > of reluctant chimeras? Replace the females with new mice (preferably in proestrus) or setting up for an IVF protocol (using survival surgery and removal of the vas deferens and epididymis from one side only - thereby maintaining the male for future breeding, and if there is a problem with the first attempt - you still have another chance at rescuing the line). The IVF scheme can also provide an index of relative fertility based on sperm concentration, morphology, and motility. Carl A. Pinkert

From TSAUNDER at hg-basic1mail.hg.med.umich.edu Wed Sep 11 14:34:42 1996 From: TSAUNDER at hg-basic1mail.hg.med.umich.edu (TSAUNDER)

Subject: Mating of chimeric mice Message-ID: <[email protected]>

David,

Our appraoch to chimeras is to give them them many mating opportunities. To wit: rotate 2 young C57BL/6 females (6-8 weeks old) through the male's cage every two weeks. This assumes that a male will have enough sperm for one fertile mating per week. Removed females are housed in pairs until it is determined whether they are gravid. By the time the third pair of females is introduced, you should have weaned pups from the first pair. Alternatively, you may wish to consider the IVF approach suggested by Carl Pinkert.

Date: Wed, 11 Sep 1996 09:02:38 +1000 To: [email protected] From: David Bowtell Subject: Mating of chimeric mice Reply-to: [email protected]

Thanks.

From jparkert at magnus.acs.ohio-state.edu Wed Sep 11 13:41:57 1996 From: jparkert at magnus.acs.ohio-state.edu (Janice Parker-Thornburg)

Subject: Mating of chimeric mice Message-ID: <[email protected]>

>We have a dozen or so chimeric mice from a KO in W9.5 cells. There is a >strong sex distortion and some of the mice are very strong. This line has >been used very successfully in a collaborating laboratory, with a high >frequency of germline transmission. The first few mice have been mated but >have not plugged C57BL/6J female over a period of about 4 weeks (these >chimeric males are now ca. 12wo). Clearly we have a way to go before we >have a problem but does anyone have any comments on 'wringing' the most out >of reluctant chimeras?

>David Bowtell

David:

When we have "reluctant" males, our favorite trick is to give them one or two superovulated females, reasoning that: 1) we are sure that the females are in heat, and 2) the "quality" of their ovulatory phase might be better (of course, this is pure conjecture. . .).

Good luck.

From carton at murray.fordham.edu Wed Sep 11 15:28:42 1996 From: carton at murray.fordham.edu (Jill Carton)

Subject: knockout mouse vector Message-ID:

Hi! I was wondering if anyone knows of a convenient vector for sale which could be used to generate a knockout transgene. I was thinking of a vector which contained the HSV-TK gene, and a neomycin resistance gene surrounded by two unique multiple cloning sites for homologous sequence insertion. Thanks.

From mlm at titus.u-strasbg.fr Thu Sep 12 12:20:39 1996 From: mlm at titus.u-strasbg.fr (Marianne LEMEUR)

Subject: NOD mice embryos Message-ID:

Has anyone an idea how to improve yields of NOD embryos. Superovulation is impossible and natural matings seem to be productive only in a narrow window.

Is there an hybrid cross which works well? We have tried B6XNOD which is not better.

Marianne Le Meur Transgenic Animal Facility IGBMC-Strasbourg, France

From pinkert at cmed.bhs.uab.edu Thu Sep 12 09:59:26 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert)

Subject: NOD zygote yields Message-ID: <19960912140427737.AAA136@cap1>

> Has anyone an idea how to improve yields of NOD embryos. Superovulation is > impossible and natural matings seem to be productive only in a narrow > window. Marianne: As per Jackson Labs, superovulation actually works better in the NOD's with older females (evidently 8-12 weeks is better than 3-4 weeks). In our first attempt this year (with approx. 8 week old females), 3 of 23 females were plugged and yielded 60 injectable eggs, of which 29 survived, and resulting in 1 stillborn and 2 resorbing fetuses. We held mice for 3 more weeks, and in our second attempt, 7 of 28 females (including some a second time) were plugged, yielding 145 injectable eggs, of which 120 survived. There were 19 liveborn pups, 18 weaned, and 4 founder transgenics. (Last year 163 eggs were injected on 2 separate days, 132 survived and were transferred, yielding 7/29 transgenics, but we went through a goodly number of mice.) AI or IVF might be more beneficial, but the time to work out the additional conditions/scheduling combined with our poor IVF yields and variable timing with some BALB/c congenics and F1's dissuaded us from going ahead in that direction. Good luck. Carl

From TSAUNDER at hg-basic1mail.hg.med.umich.edu Fri Sep 13 13:31:13 1996 From: TSAUNDER at hg-basic1mail.hg.med.umich.edu (TSAUNDER)

Subject: Forwarded: Ovulation Model Message-ID: <[email protected]>

This message posted on behalf of Jennifer Bowen, Please direct replies to [email protected] -Thanks, Thom Saunders I am interested in using a mouse model consisting of immature mice stimulated to ovulate a normal or near-normal number of follicles (ie not superovulated) and then mated to vasectomized males to induce pseudopregnancy. I am designing this model for future use in a mutant which is on a C57BL/6J X C3HB/FeJ background so will probably use C57BL/6 animals. I would appreciate any information on what sort of hormone dose/age combination will result in a normal number of ovulations in immature females of this or similar strains.

Jennifer Bowen ([email protected])

From mlm at titus.u-strasbg.fr Fri Sep 13 17:34:49 1996 From: mlm at titus.u-strasbg.fr (Marianne LEMEUR)

Subject: sending frozen embryos Message-ID:

I would like to send frozen embryos all over the world instead of sending living mice, which is very time consuming to get all the formalities together.

What are the conditions to do that, and were is it possible to buy the container. In France, our transportation company told us that the sender is personnally responsible in case of an accident with liquid nitrogen during the flight!

Marianne Le Meur IGBMC Strasbourg-France

From ucympek at ucl.ac.uk Fri Sep 13 16:47:32 1996 From: ucympek at ucl.ac.uk (Peter Koder)

Subject: sending frozen embryos Message-ID:

Hello Marianne

For a UK> recipient an Import licence is required from the Ministry of Agriculture tel (0)181 330 8178 under the Importation of Embryos, Ova and Semen order. Also the permission of a Home Office Inspector under the UK Animals(Scientific Procedures) Act 1986 The importing institute Genetically Modified Organism (GMO) safety committee has also to be informed inorder to accept them under the Environmental Protection Act (Contained Use) Regulations.

I hope this helps, Peter

I would like to send frozen embryos all over the world instead of sending >living mice, which is very time consuming to get all the formalities >together. > >What are the conditions to do that, and were is it possible to buy the >container. >In France, our transportation company told us that the sender is >personnally responsible in case of an accident with liquid nitrogen during >the flight! > >Marianne Le Meur >IGBMC >Strasbourg-France > > > > > >

Peter C Koder BVMS MSc MRCVS UCL Biological Services E-MAIL: [email protected] University College London TEL:+44(0)171 391 1309 Malet Place FAX:+44(0)171 380 7837 LONDON, WC1E 6BT UK

From pinkert at cmed.bhs.uab.edu Fri Sep 13 11:55:51 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert)

Subject: sending frozen embryos Message-ID: <19960913160057705.AAA159@cap1>

> Date: Fri, 13 Sep 1996 16:34:49 +0000 > To: [email protected] > From: [email protected] (Marianne LEMEUR) > Subject: sending frozen embryos > Reply-to: [email protected]

> I would like to send frozen embryos all over the world instead of sending > living mice, which is very time consuming to get all the formalities > together. > > What are the conditions to do that, and were is it possible to buy the > container. > In France, our transportation company told us that the sender is > personnally responsible in case of an accident with liquid nitrogen during > the flight! You may want to check with Dr. Glenn Monastersky regarding shipment of ova and related considerations. He was using the liquid nitrogen shipping containers that contained an absorbant material - it maintained temperature, yet there was no LN2 sloshing around at all. (what they used I believe was similar to the Arctic Express from Thermolyne (model CY50915) that keeps temperature for 11 days) . Glenn can be contacted at [email protected]. Carl

From andreja at ariel.ucs.unimelb.edu.au Sat Sep 14 01:09:28 1996 From: andreja at ariel.ucs.unimelb.edu.au (Andreja Pirkmaier)

Subject: Caesarean sections Message-ID: <[email protected]>

Has anyone any experience with injecting mice with hormones( Day, Dose) few Days prior doing caesarean section to avoid any early delivery. I know that the gestation period for the transgenic line I am interested in is 19 Days. I planed to do sections on Day 18 p. c..

Thanks Andreja Pirkmaier Peter MacCallum Cancer Institute [email protected]

From tonyjames at cuhk.edu.hk Sat Sep 14 02:43:18 1996 From: tonyjames at cuhk.edu.hk (A E James)

Subject: Caesarean sections Message-ID: <[email protected]>

> Has anyone any experience with injecting mice with hormones( Day, >Dose) few Days prior doing caesarean section to avoid any early delivery. > I know that the gestation period for the transgenic line I am >interested in is 19 Days. I planed to do sections on Day 18 p. c.. > >Thanks Andreja Pirkmaier >Peter MacCallum Cancer Institute >[email protected] > > > >Try Catherine O'Brien at WEHI or her senior technician at WEHI Kew...her name is Therese Johns. Any way my protocol is progesterone for three days before the expected birth. In pregnant rats the figure quoted is 20 mg per day for a heavy preg. rat. Other figures I have seen for mice are 1.5 to 2.5 mg per day per mouse for the three days prior to csaerian delivery. Hope this helps Tony James Tony James BVSc MACVSc Director Laboratory Animal Unit Chinese University of Hong Kong Shatin, New Territories, Hong Kong ph: 852 2609 6862 fx: 852 2603 5723 email: [email protected] ...... "nothing great was achieved without enthusiasm." R. W. Emerson

From pinkert at cmed.bhs.uab.edu Sat Sep 14 09:48:18 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert)

Subject: Caesarean sections Message-ID: <19960914135340470.AAA136@cap1>

> Has anyone any experience with injecting mice with hormones( Day, > Dose) few Days prior doing caesarean section to avoid any early delivery. > I know that the gestation period for the transgenic line I am > interested in is 19 Days. I planed to do sections on Day 18 p. c.. If the line uniformly delivers on d19 and the pups are robust, then C-sections on d18 should likely work out well without the additional hormone therapy.

From mlm at titus.u-strasbg.fr Tue Sep 17 16:19:26 1996 From: mlm at titus.u-strasbg.fr (Marianne LEMEUR)

Subject: zygote yields Message-ID:

Thanks to those who gave the solution for NOD mice zygote yield! More generally it seems that C57Bl6 is the only inbred strain which can be superovulated in the same way as an F1 hybrid. Other inbred strains like FVB/N or outbred strains like CD1 do not give better yields than can be achieved with a natural mating.Is that correct? Marianne Le Meur From jparkert at magnus.acs.ohio-state.edu Tue Sep 17 15:45:24 1996 From: jparkert at magnus.acs.ohio-state.edu (Janice Parker-Thornburg)

Subject: zygote yields Message-ID: <[email protected]>

Marianne:

This is not correct. I have superovulated FVB/N for microinjection, as well as for general harvesting of mouse embryos. If done in the correct time frame, one can get 8-10 more eggs (embryos) per female than with natural matings. I also know that people using CD1's for embryo harvesting will superovulate them, presumably because they get a better yield. If you find that your yield does not improve with superovulation, it is likely that your mouse-age or timespan for the hormone injections needs to be optimized.

From landel at medsci.udel.edu Tue Sep 17 16:37:11 1996 From: landel at medsci.udel.edu (Carlisle Landel)

Subject: zygote yields Message-ID: <[email protected]>

Marianne,

No, this isn't correct! I superovulate FVB/N routinely, and I'm pretty sure that colleagues of mine in the past have done it to CD1's, too, though I vaguely recall there was some trick with dosage or timing or something (but I may be wrong--this was many years ago).

Check your hormones for proper concentration, and check that the light controls in your facility are properly set, and make sure that you are injecting at or very near the middle of your light cycle.

Regards,

Carlisle Landel Dept. of Clinical Science A.I. duPont Institute PO Box 269 Wilmington DE 19899 (302) 651-6873 [email protected] http://mdblmac.medsci.udel.edu/WEB/MDBL/Carlisle.html

From pinkert at cmed.bhs.uab.edu Tue Sep 17 10:23:08 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert)

Subject: zygote yields Message-ID: <19960917142758010.AAB158@cap1>

> More generally it seems that C57Bl6 is the only inbred strain which can be > superovulated in the same way as an F1 hybrid. I'm not sure what is meant here, but other inbred strains can be utilized, using the same general scheme and sometimes jogging the schedule a bit.

> Other inbred strains like FVB/N or outbred strains like CD1 do not give > better yields than can be achieved with a natural mating.Is that correct? It's a matter of degree, but I disagree with the last statement (unless you mean that hybrids provide better net yields than either inbreds and outbreds, which is true). In some labs, natural matings facilitate a specific routine. However, there are significant economies-of-scale when superovulation is employed (in relation to zygote yields and needed labor/space/etc.) when using the strains that you have mentioned.

From browng at medicine.wustl.edu Tue Sep 17 19:03:45 1996 From: browng at medicine.wustl.edu (Gary Brown) Subject: zygote yields In-Reply-To: Message-ID:

On Tue, 17 Sep 1996, Marianne LEMEUR wrote:

> Thanks to those who gave the solution for NOD mice zygote yield! > More generally it seems that C57Bl6 is the only inbred strain which can be > superovulated in the same way as an F1 hybrid. > Other inbred strains like FVB/N or outbred strains like CD1 do not give > better yields than can be achieved with a natural mating.Is that correct? > Marianne Le Meur > > > > Not so. I used FVB/N strain mice for 3 1/2 years and superovulated with (in my opinion) a high degree of success. Since it's a larger mouse than B6C3F1 hybrids, we used 7.5IU hormones rather than 5IU, and a 47 hr gap between PMSG and HCG. Typical yields were 25-30 good (non fragmented) eggs per mouse. Can't say I've had experience with other strains other than C57Bl/6 and B6C3F1 hybrids. Actually, I lie. 129SvJ mice are dismal for superovulation - if anyone can give me a way to do these reproducibly I'll get you a beer (or three)!

From carboni at valoritech.com Tue Sep 17 20:25:09 1996 From: carboni at valoritech.com (Nick Carboni)

Subject: Wood's coculture technique Message-ID: <[email protected]>

Hi everybody,

Short message to know if one of you is currently working or has tried to work on wood's coculture technique.

I have spoke with some people, and it appears that it is rather difficult to acheive. Has anyone any experience to share with me ??

Thanks in advance,

Nick ------Nick Carboni, VP Valoritech 1253, McGill College Av., room 620 Montreal, Quebec, Canada H3B 2Y5 Tel.: (514) 866-8271 / Fax: (514) 866-8272 E-Mail: [email protected]

From browng at medicine.wustl.edu Wed Sep 18 13:26:12 1996 From: browng at medicine.wustl.edu (Gary Brown)

Subject: Wood's coculture technique In-Reply-To: <[email protected]> Message-ID:

On Tue, 17 Sep 1996, Nick Carboni wrote:

> Hi everybody, > > > Short message to know if one of you is currently working or has tried to > work on wood's coculture technique. > > I have spoke with some people, and it appears that it is rather difficult to > acheive. > > Has anyone any experience to share with me ?? > > Thanks in advance, > > Nick > Hi !

I tried Wood's co-culture technique around 3 years ago, but only had 1 recorded birth in 16 implanted mothers, and this pup was malformed. We were unable to determine the efficacy of the technique as the ES cell line that we used was obtained from another laboratory, and both their technique and the cell line itself were called into question. However, here's what I can tell you..

1. We found that the ES cells did not adhere to the de - ZP'd morulae / octoploid / tetraploid embryos as well as we thought the paper indicated.

2. Our understanding is that the technique's success is very ES cell dependent.

3. Our understanding is that the live birth rate is very low, making this a system better suited to in utero developmental studies.

Since this time, I have tried to find the time to explore non-injection methods of chimera production (with little success on the time front!). I have only managed to complete a pilot study of Nagy / Rossant's "sandwich" method where I was successful in fusing 2 morulae together in culture overnight in custom made dimples. This effort would have been more instructive if I had had access to some ES cells to "sandwich", but the fusions had a >90% success rate. For the reference on this methodology, do a Medline search on the authors above or check out the E-newsletter archive on my homepage for August / September.

Hope this is instructive! :-)

From ldeg at midway.uchicago.edu Thu Sep 19 02:28:26 1996 From: ldeg at midway.uchicago.edu (Linda Degenstein)

Subject: ES Cells from Balb C mice Message-ID: <[email protected]>

We are looking for ES Cells (germline proven) that are made from Balb C strain mice. Does anyone know if these are available?

From khelmuth at facstaff.wisc.edu Tue Sep 24 20:28:52 1996 From: khelmuth at facstaff.wisc.edu ([email protected])

Subject: noise problems Message-ID: Hi everyone:

I am looking for documentation that excess noise can affect the breeding and production of embryos from mice. Does anyone know of published work that looked at environmental affects on a mouse colony? Any citations would be appreciated.

From mlp at informatics.jax.org Tue Sep 24 22:20:46 1996 From: mlp at informatics.jax.org (Moyha Lennon-Pierce)

Subject: Noise/reproduction Message-ID:

The following references might be useful.

I searched the Strain Characteristics Catalog in development at The Jackson Lab and did a quick search in NLM. Not much at all in the literature, but the following references may be useful.

Nawrot PS et al. Embryotoxicity of various noise stimuli in the mouse. Teratology 1980; 22(3):279-89. CF-1

Zakem HB, Alliston CW. The effects of noise level and elevated ambient temperatures upon selected reproductive traits in female Swiss-Webster mice. Lab Anim Sci 1974; 24:469-75. Strain SWR

Moyha Lennon-Pierce MGD Informatics [email protected]

>Hi everyone: > >I am looking for documentation that excess noise can affect the breeding >and production of embryos from mice. Does anyone know of published work >that looked at environmental affects on a mouse colony? Any citations >would be appreciated. > >Kathy > >Kathy Krentz Helmuth >Transgenic Facility >425 Henry Mall >Room 2210 Biotechnology Center >Madison, WI 53706 >(608) 265-2801 >email:[email protected]

From landel at medsci.udel.edu Tue Sep 24 22:13:36 1996 From: landel at medsci.udel.edu (Carlisle Landel)

Subject: Noise/reproduction Message-ID: <[email protected]>

Kathy,

I saw a poster at a meeting a couple of weeks ago where they were using one of those ultrasonic mouse repellers to increase the percentage of embryo loss in CBAxDBA/2 model. If you want more info, I'll see if I can point you towards the authors.

Regards,

Carlisle Landel Dept. of Clinical Science duPont Hospital for Children PO Box 269 Wilmington DE 19899 (302) 651-6873 [email protected] http://mdblmac.medsci.udel.edu/WEB/MDBL/Carlisle.html

From Roger.Leemann at imr.psi.ch Wed Sep 25 15:40:47 1996 From: Roger.Leemann at imr.psi.ch (Roger C. Leemann)

Subject: noise problems Message-ID: <[email protected]>

Kathy Krentz Helmuth ([email protected]) wrote:

>I am looking for documentation that excess noise can affect the breeding >and production of embryos from mice. Kathy

Sorry, no literature, but I can tell you how we do it. In our animal housing facility there is always a radio playing at a low volume during the daylight cycle (maybe this can not be referred to as "excessive" noise.) I don't know whether or how this affects the mice, positive or negative. With non-superovulated B6C3F1 mice I can expect 15% to 25% plugs and 7 to 12 two-cell embryos/mouse. How does this compare to other labs?

1996 October

From pasceri at sickkids.on.ca Tue Oct 1 17:34:58 1996 From: pasceri at sickkids.on.ca (Peter Pasceri) Date: Wed Aug 20 11:52:53 2003 Subject: Cryopreservation Message-ID:

Hi all!

I am looking for suppliers of mouse embryo freezing kits. Can you help?

Thanks,

-- Peter Pasceri [email protected] Genetics Research Phone: (416) 813-4205 The Hospital for Sick Children Toronto, Ontario,Canada

From p.sobieszczuk at ic.ac.uk Thu Oct 3 12:37:29 1996 From: p.sobieszczuk at ic.ac.uk (Peter Sobieszczuk) Date: Wed Aug 20 11:52:53 2003 Subject: Transgenic mice/Transplantation (fwd) Message-ID:

Sorry, if it is a duplication for some of you. Peter (your listowner)

>Hello everyone, > >Any advise/suggestions on the following questions would be greatly >appreciated. > >Thank you > >Anna V. Anagnostopoulos >TBASE >Division of Biomedical Information Sciences >The Johns Hopkins University School of Medicine >[email protected] > >------> >I am working with mice, doing Bone Marrow and Peripheral Blood >Tranplantation experiments, at the Department of Immunogenetics of the New >York Blood Center. We are interested in developing a syngeneic mouse >transplantation model. We would like to use as donors, animals carrying a >transgene, that its product is not lethal or significantly affecting the >recipients (especially hematologically), and also is easily detectable >post-transplant. For example, mice transgenic for a human gene, encoding >for a human antibody, would be a good model. The idea would be to show >that the donor cells (positive for the transgene) engraft, and the >transgene is functional (we can detect the product) after transplant. The >mouse strains I am working with at the present time are C57BL/6, A/J and >B6A (C57BL/6 X A/J) (all obtained from Jackson Labs). It would help >tremendously if the transgenic animals are of the same background. Do you >have any suggestions? >

From mangia at uniroma1.it Fri Oct 4 13:46:36 1996 From: mangia at uniroma1.it (Franco Mangia) Date: Wed Aug 20 11:52:53 2003 Subject: Transgenic mice/Transplantation (fwd) Message-ID: > >We are interested in developing a syngeneic mouse >>transplantation model. We would like to use as donors, animals carrying a >>transgene, that its product is not lethal or significantly affecting the >>recipients (especially hematologically), and also is easily detectable >>post-transplant. For example, mice transgenic for a human gene, encoding >>for a human antibody, would be a good model. The idea would be to show >>that the donor cells (positive for the transgene) engraft, and the >>transgene is functional (we can detect the product) after transplant. The >>mouse strains I am working with at the present time are C57BL/6, A/J and >>B6A (C57BL/6 X A/J) (all obtained from Jackson Labs). It would help >>tremendously if the transgenic animals are of the same background. Do you >>have any suggestions? >> >Anna V. Anagnostopoulos >TBASE >Division of Biomedical Information Sciences >The Johns Hopkins University School of Medicine >[email protected]

I think that the ROSA26 transgenic mice, developed by Phil Soriano, are just what you need for labeling engrafts at a single cell level. These mice carry a lacZ transgene directed by a ubiquitous promoter active in all tissues, without apparently affecting cell viability, since these mice are fine. These mice can be obtained from the Jackson Labs either as B6,129 F1 (the original Soriano's hybrid strain) or with a pure C57BL/6J background. Complete names of these mice are: "B6,129-TgR(ROSA26)26 Sor", and "C57Bl/6J-TgR(ROSA26)26 Sor". I am also very interested in obtaining an ES cell line(s) from these mice. Is anybody aware whether these ES lines are being developed in some lab? Also, does anybody know whether ROSA26 mice with nuclearly targeted beta-gal have been produced? Franco

Franco Mangia Laboratory of General Biology Department of Psychology University La Sapienza of Rome c/o Institute of Histology and General Embryology Via Borelli, 50 00161 Rome, Italy tel: 39-6-4976-8103 FAX: 39-6-4976-8099 E-mail: [email protected]

From p.sobieszczuk at ic.ac.uk Sat Oct 5 19:19:10 1996 From: p.sobieszczuk at ic.ac.uk (Peter Sobieszczuk) Date: Wed Aug 20 11:52:53 2003 Subject: "TgList Backstage" (admin. news *2) ARCHIVES & TBASE Message-ID:

Dear Transgenic Subscribers,

Greetings to newcomers,

ARCHIVES Charmaine Foltz asked me some time ago: >I was wondering if the information on the list will be retrievable >(i.e., will there be a data base that we can search archived files). very much so, send the message: index transgenic-list to listserver at [email protected] and you shall receive an index of monthly files, something like:

>>>> index transgenic-list total 241 -rw-rw---- 1 daemon 10364 Jul 29 16:38 transgenic- list.archive.9607 -rw-rw---- 1 majordom 146666 Aug 29 21:36 transgenic- list.archive.9608 -rw-rw---- 1 majordom 71873 Sep 25 15:31 transgenic- list.archive.9609 -rw-rw---- 1 majordom 6715 Oct 4 12:46 transgenic- list.archive.9610 >>>> where July entries file name is "transgenic-list.archive.9607"

If you then send: get transgenic-list transgenic-list.archive.9607 you should receive all postings for that month (in this case, July) as individual e-mail messages.

If you have access to world wide web, (WWW) you can explore archives with the luxury of selecting individual entries etc., using your favourite browser, Netscape, Mosaic or Explorer.

The URL address is: http://www.lists.ic.ac.uk/hypermail/transgenic-list I'm afraid however that even with the browser, files are not searchable (at this stage anyway), although subject, (which should be the topic of any message) is clearly visible.

TBASE Anna Anagnostopoulos who is co-ordinating TBASE data base site sent this introduction (with minor editorial changes P.S.):

------BEGINNING OF TEXT

TBASE, the Transgenic/Targeted Mutation Database, is an attempt to organize information on transgenic animals and targeted mutations generated, and analyzed worldwide. TBASE is available on the WWW http://www.gdb.org/Dan/tbase/tbase.html and can be searched either quite broadly across the entire database (the 'blunt object' approach) or quite specifically with the use of 28 distinct searchable fields (the 'scalpel and tweezers' approach). TBASE contains a wealth of information about each animal in the database. A selection of the fields present in the databases are: line name, genus, DNA construct description, host background, line source, method used , transgene/targeted allele expression, lethality, phenotype, handling, contact information, full citation and abstracts of the papers related to the creation of a line.

Owing to the wide use of homologous recombination in ES cells, TBASE initially focused on maintaining a comprehensive coverage of all targeted mutations reported to date. Moreover, it focused on the mouse as the predominant mammalian model. Specific emphasis is still placed on cataloging novel knockout mice, which have defined genetic modifications, with an extensive detailed description of their resulting mutant phenotypes. In the future, the focus of TBASE will expand to cover all animals resulting from transgenesis or targeted mutation, regardless of species. Additionally, TBASE plans to incorporate 'transgenic knockouts', that is, knockout animals that have subsequently served as recipients for transgenes.

TBASE should be viewed as a dynamic database with a potentially unlimited number of entries, expected to grow considerably as more experimental methods and lines are generated. Data acquisition is primarily effected by active literature scanning and manual data entry, as well as processing of direct submission forms.

Literature scanning refers to direct data extraction from the scientific literature through regular examination of over 20 journals. These journals have been statistically identified as periodically stable sources of information on transgenesis and gene targeting. Active literature scanning has, so far, been the primary mode of data accumulation, and has ensured that the database faithfully reflects the general direction of related research. It is expected to remain a significant component of the data acquisition tactics, even as other methods of data acquisition become available (see below). In an effort to avoid missing important data, keyword-related references from additional journals are also screened by the use of Medline and Current Contents.

In the future, data that are not manually entered by TBASE staff through the literature scanning process, are expected to arrive either as direct paper or as electronic submissions. Although over 98% of the data have been entered manually so far, direct paper submissions are expected to increase as TBASE becomes fully adopted by the scientific community. In addition, as electronic submission tools become available, more research groups will be prepared to commit resources to the production of data deposited into TBASE. Electronic submission tools and their error-checking capabilities will ensure efficient and timely deposition of the data.

END OF TEXT ------

STATISTIC: There are approx. 240 members from at least 18 countries registered now, and I'm starting to think aren't we growing too big, read to much to read.

Regards,

Yours sincerely,

Listowner

______

Dr Peter Sobieszczuk Imperial College School of Medicine at St. Mary's tel: 0171 594 3784 Biochemistry and Molecular Genetics tel: 0171 594 3799 Norfolk Place fax: 0171 706 3272 London W2 1PG, U.K. e-mail: [email protected] ______

From khelmuth at facstaff.wisc.edu Wed Oct 9 10:28:47 1996 From: khelmuth at facstaff.wisc.edu ([email protected]) Date: Wed Aug 20 11:52:53 2003 Subject: fire codes within animal colony Message-ID:

Hello everyone-

I face a delemma within our facility and am hoping someone can help me out. We recently moved our rat and mouse colonies into a new building. Everything seems to be going well except for one thing. We experienced the first fire drill and realized that there are 3 fire alarms within our 5ft. wide clean hallway and 3 fire alarms within our 5ft. wide dirty hallway. They are extremely loud and potentially problematic for our animals. Our building manager agrees that this seems extremely overdone and wishes to get some of the alarms diconnected. However, the state fire code regulator feels that with the background noises of the cage washer, movement of carts and racks, etc. that these extra fire alarms are necessary. He will only follow suit if I can come up with evidence that not only does it affect the well-being of the animals but that other facilities have a small number of alarms. I have obtained documentation of loud noises and it's effect on breeding, embryo production, etc. Can someone help me out with regulations regarding fire alarms within your animal facility?

Any suggestion on how to deal with this would be beneficial. Thanks

Kathy Krentz Helmuth

Kathy Krentz Helmuth Transgenic Facility 425 Henry Mall Room 2210 Biotechnology Center Madison, WI 53706 (608) 265-2801 email:[email protected] From landel at medsci.udel.edu Wed Oct 9 11:44:03 1996 From: landel at medsci.udel.edu (Carlisle Landel) Date: Wed Aug 20 11:52:53 2003 Subject: fire codes within animal colony Message-ID: <[email protected]>

I'm curious--how often do these alarms go off? Maybe instead you could get them to absolve you from frequent fire drills.

Carlisle Landel

>From [email protected] Wed Oct 9 10:35:44 1996 >Mime-Version: 1.0 >Content-Type: text/plain; charset="us-ascii" >To: [email protected] >From: [email protected] >Subject: fire codes within animal colony >Sender: [email protected] >Precedence: bulk >Reply-To: [email protected] > >Hello everyone- > >I face a delemma within our facility and am hoping someone can help me out. >We recently moved our rat and mouse colonies into a new building. >Everything seems to be going well except for one thing. We experienced the >first fire drill and realized that there are 3 fire alarms within our 5ft. >wide clean hallway and 3 fire alarms within our 5ft. wide dirty hallway. >They are extremely loud and potentially problematic for our animals. Our >building manager agrees that this seems extremely overdone and wishes to >get some of the alarms diconnected. However, the state fire code regulator >feels that with the background noises of the cage washer, movement of carts >and racks, etc. that these extra fire alarms are necessary. He will only >follow suit if I can come up with evidence that not only does it affect the >well-being of the animals but that other facilities have a small number of >alarms. I have obtained documentation of loud noises and it's effect on >breeding, embryo production, etc. Can someone help me out with regulations >regarding fire alarms within your animal facility? > >Any suggestion on how to deal with this would be beneficial. >Thanks > >Kathy Krentz Helmuth > >Kathy Krentz Helmuth >Transgenic Facility >425 Henry Mall >Room 2210 Biotechnology Center >Madison, WI 53706 >(608) 265-2801 >email:[email protected] > > > > >

From duffyh at war.wyeth.com Wed Oct 9 15:45:06 1996 From: duffyh at war.wyeth.com (Heidi Duffy) Date: Wed Aug 20 11:52:53 2003 Subject: fire codes within animal colony -Reply Message-ID:

I used to work in animal facility that used flashing lights instead of audible alarms within the animal facility. The audible alarms were only in the office areas of the facility. (Our phones within the facility were set up the same way, including the cagewash areas). Designated techs were assigned to check all animal rooms for personnel evacuation before leaving the buildings. It worked out very well.

Heidi Duffy email:[email protected]

From david_gask at Merck.Com Thu Oct 10 13:47:00 1996 From: david_gask at Merck.Com (David Gask) Date: Wed Aug 20 11:52:53 2003 Subject: Fire Alarm problems-Kathy Krentz Helmuth Message-ID: <199610101150.HAA21899@igw2>

Kathy, Why don't you investigate the use of ' SILENTONE ALARMS' as an alternative. These are used widely in the UK and were developed to prevent problems for the animals caused by more conventional sounders. They produce a loud noise suitable for use as an alarm in the event of fire, but at the same time the frequency is below the threshold frequency, or outside the most sensitive range for most commonly used laboratory animals. If you require the address of a supplier I can send it to you. All our animal facility fire alarms are of this type.

Hope this helps

Dave Gask Operations Coordinator Merck Sharp & Dohme Neuroscience Research Centre Terlings Park Eastwick Road Harlow, Essex CM20 2QR England Tel 01279 440220 e-mail [email protected]

Hello everyone-

From Roger.Leemann at imr.psi.ch Thu Oct 10 16:21:53 1996 From: Roger.Leemann at imr.psi.ch (Roger C. Leemann) Date: Wed Aug 20 11:52:53 2003 Subject: Rodent-research list Message-ID:

Since I'm working with mice but do not use transgenic animals, I subscribed to the rodent-research list about a month ago. Majordomo told me, that it had forwarded my request to the list owner for approval but I haven't got any response since. Maybe it's not a list for every Jack and Joe (or was it Jack and Jill?), or maybe the list owner is too busy, or on vacation, whatever. I don't mind if they don't want me, but ones likes to know... Anybody in this list who has a clue why the rodent-research list seems dead?

From p.sobieszczuk at ic.ac.uk Thu Oct 10 17:55:30 1996 From: p.sobieszczuk at ic.ac.uk (Peter Sobieszczuk) Date: Wed Aug 20 11:52:53 2003 Subject: Rodent-research list & "TgL Backstage"(admin.news*3) Message-ID:

Dear Roger,

>Since I'm working with mice but do not use transgenic animals, I subscribed >to the rodent-research list about a month ago. Majordomo told me, that it I presume listserver "majordomo" at Caltech, and not "ic.ac.uk" at Imperial College?

>Maybe it's not a list for every Jack and Joe (or was it Jack and Jill?), or Do not take it personnally, it's just a computer. >maybe the list owner is too busy, or on vacation, whatever. I don't mind if >they don't want me, but ones likes to know...

Sorry, I still do not know what exactly happened, Eric Mercer had some technical problems supporting "rodent-research" list, 3 or 4 months ago. A few people has tried to contact him, at the end, this list was created (see the introduction to the list available also by sending "info transgenic-list" message to "[email protected]" address).

You have been with us (transgenic-list) for the month now and probably noticed that the list is un-moderated and has no restriction on the content. Transgenic - is just a name on a server, and the list a quick way to exchange information.

I would not like to start a discussion on definition of transgenesis, but would imagine that for the active researchers in the field it would cover quite a bit of biology, from molecular to developmental, with the animal models not exclussively restricted to rodents.

So it is not the name but participants who will take the discussion in any particular direction, including yourself.

Who would you like to see on the list Roger, or should I ask everybody:

Who else do you think we should invite to the list?

Best regards,

Sincerely,

Peter - your listowner

From s.tan at anatomy.unimelb.edu.au Tue Oct 15 16:09:53 1996 From: s.tan at anatomy.unimelb.edu.au (S.Tan) Date: Wed Aug 20 11:52:53 2003 Subject: Fostering baby mice Message-ID:

I am grafting pieces of neural tissue into the cortex of newborn mice but find that the mother keeps eating the babies. The wound is closed with a single silk suture and this appears to attract the attention of mum. The mother's strain is C57/BL6 x DBA. Are there less aggressive strains of mums out there? Thanks. From a.annala at ucl.ac.uk Tue Oct 15 06:51:24 1996 From: a.annala at ucl.ac.uk (Dr. Alexander J. Annala) Date: Wed Aug 20 11:52:53 2003 Subject: Fostering baby mice Message-ID:

>I am grafting pieces of neural tissue into the cortex of newborn mice but >find that the mother keeps eating the babies. The wound is closed with a >single silk suture and this appears to attract the attention of mum. The >mother's strain is C57/BL6 x DBA. Are there less aggressive strains of >mums out there? Thanks.

Most F1's make very good mothers. Your C57BL/6 x DBA are F1 mice. We use C57BL/6 x CBA and C57BL/10 x CBA with good results.

Do you take precaution to avoid leaving scent on operated baby mice? Some people roll babies in mom's urine before returning them to the nest. Others treat mom's nose with alcohol to keep her from smelling any differences between babies for awhile. This might be a scent issue rather than an exposed suture problem.

Internalize suture might be difficult in baby mice. Maybe you could make your incision somewhere away from your skull opening, slide the skin opening over the area you want to open bone, do your work, slide the skin back, and maybe use one of the tissue adhesives which are pretty available around medical schools and operating theatres to close the wound. This would avoid leaving exposed suture for the mom to chew on.

From fobo at ltk.unizh.ch Tue Oct 15 08:22:40 1996 From: fobo at ltk.unizh.ch (Frank Bootz) Date: Wed Aug 20 11:52:53 2003 Subject: Fostering baby mice Message-ID: <[email protected]>

Dear colleaque,

I have done thymectomy in newborn mouse. The wound was also closed with a single U-suture in the neck. The pups awake within 5 minutes after methofane (methoxyflurane) anesthesia and brought back to the mother immediately. We don't anesthetize the mother, contrary the most papers describing this procedure. In this case the mother's strain is C57/Bl/6 x PO. If we can choose the strain, we normally use Zur:ICR original Charles River swiss mice CD-1. They are mice with excellent mother attributes. Best regards,

Frank Frank Bootz Institute of Laboratory Animal Science University of Zurich

Winterthurerstrasse 190 phone: +41 1 257 54 55 8057 Zurich fax: +41 1 257 57 03 Switzerland e-mail [email protected]

From GWOLFF at NCTR.FDA.GOV Tue Oct 15 08:44:51 1996 From: GWOLFF at NCTR.FDA.GOV (George L. Wolff) Date: Wed Aug 20 11:52:53 2003 Subject: Fostering baby mice Message-ID: <[email protected]>

Zur:ICR cannot be descended from CD-1 mice - or the strain designation is incorrect. Let me know if you'd like more details. George L. Wolff

Tel: (501) 543-7522 FAX: (501) 543-7635/7662 NCTR HFT-140 3900 NCTR Road Jefferson, AR 72079-9502

From gotto at leland.Stanford.EDU Wed Oct 16 13:03:00 1996 From: gotto at leland.Stanford.EDU (Glen Otto) Date: Wed Aug 20 11:52:53 2003 Subject: Fostering baby mice In-Reply-To: <[email protected]> Message-ID:

A non-text attachment was scrubbed... Name: not available Type: text/enriched Size: 577 bytes Desc: not available Url : http://mailman.ic.ac.uk/mailman/private/transgenic- list/attachments/19961016/3cf223e6/attachment.bin From a.annala at ucl.ac.uk Wed Oct 16 22:50:59 1996 From: a.annala at ucl.ac.uk (Dr. Alexander J. Annala) Date: Wed Aug 20 11:52:53 2003 Subject: Fostering baby mice -- Failure to thrive? Message-ID: What can be done to assist baby mice in distress after birth? Some chimeras with neuroreceptor channel mutations experience difficulty feeding themselves -- and sometimes even suckling milk from mother.

We have in the past injected approx 20% weight of animal with Ringers Lactate (Hartman's Solution). However there must be more aggressive therapy which would facilitate survival of distressed baby mice.

We have also used incubator -- with reservations about the possibility of dehydration from elevated environmental temperature.

Thanks,

Alexander J. Annala, Ph.D. Senior Research Fellow Laboratory for Molecular Pharmacology University College London

From carton at murray.fordham.edu Wed Oct 16 19:44:40 1996 From: carton at murray.fordham.edu (Jill Carton) Date: Wed Aug 20 11:52:53 2003 Subject: knockout mouse construct problems Message-ID:

We are working on generating several knockout mice and have been successful in producing transgenes for some of our genes, but we have a fairly consistent problem with the final ligation step of assembling the construct. When we PCR our ligation reaction to see if the DNA pieces have ligated we do get the appropriate product however analysis of the clones by miniprep show no positive clones. So we are suspecting that the problem is in the uptake or growth of the construct in the bacterial cells. Although, each fragment of the transgene can be propagated in the bacteria. At this stage we are working with a very large piece of DNA (~12KB)- and perhaps this will lower the ligation frequency-but we have generated larger plasmids with little problem. We are using JM109 or DH5a as our competent cells (chemically competent). Any suggestions at all would be greatly appreciated!

Jill Carton Dept. Biological Sciences Fordham University Larkin Hall, rm 160 Bronx, NY 10458 718-817-3652 718-817-3645 (fax) [email protected] From p.sobieszczuk at ic.ac.uk Thu Oct 17 16:23:53 1996 From: p.sobieszczuk at ic.ac.uk (Peter Sobieszczuk) Date: Wed Aug 20 11:52:53 2003 Subject: Nod/Scid mice Message-ID:

Forwarded from [Anna Anagnostopoulos ]

>Hello everyone, > >Dr. Sally J. Cutler (Department of Medical Microbiology, Charing Cross >Hospital, London. Telephone:+ 44181 846 7570) is looking for sources of >Nod/Scid mice, preferably in the UK. Any suggestions on where to look? > >Thank so much > >~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ >Anna V. Anagnostopoulos, Ph.D. >Division of Biomedical Information Sciences >The Johns Hopkins University School of Medicine >2024 E. Monument Street >Baltimore, MD 21205-2236 >tel: (410) 614-3226 >fax: (410) 614-0434 >e-mail: [email protected]

From james at icr.ac.uk Thu Oct 17 18:00:12 1996 From: james at icr.ac.uk (james wallace) Date: Wed Aug 20 11:52:53 2003 Subject: Nod/Scid mice Message-ID:

> >Dr. Sally J. Cutler (Department of Medical Microbiology, Charing Cross > >Hospital, London. Telephone:+ 44181 846 7570) is looking for sources of > >Nod/Scid mice, preferably in the UK. Any suggestions on where to look? > > > >Thank so much > > > >~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ > >Anna V. Anagnostopoulos, Ph.D. > >Division of Biomedical Information Sciences > >The Johns Hopkins University School of Medicine > >2024 E. Monument Street > >Baltimore, MD 21205-2236 > >tel: (410) 614-3226 > >fax: (410) 614-0434 > >e-mail: [email protected] > > Hi, Tell Sally to give me a call as we have a colony of Nod\Scids here at ICR in the UK.

Best Wishes

Jim Wallace Institute of Cancer Research The McElwain Laboratories 15 Cotswold Rd Belmont Sutton Surrey SM2 5NG England tel: 0181 643 8901 Ext 4638 Fax: 0181 770 1395 e_mail [email protected] > >

From anna at screams.gdb.org Thu Oct 17 14:18:07 1996 From: anna at screams.gdb.org (Anna Anagnostopoulos) Date: Wed Aug 20 11:52:53 2003 Subject: Available enhancer traps Message-ID:

Hello again,

If any of you is interested in Joe Miano's enhancer traps or knows of a bulletin where he can have the pictures posted please let me know.

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ Anna V. Anagnostopoulos, Ph.D. Division of Biomedical Information Sciences The Johns Hopkins University School of Medicine 2024 E. Monument Street Baltimore, MD 21205-2236 tel: (410) 614-3226 fax: (410) 614-0434 e-mail: [email protected]

------Forwarded message ------Date: Wed, 16 Oct 1996 16:36:44 -0500 From: Joe Miano To: [email protected] Subject: enhancer trap

We do transgenics and I have two questions:

(1) Would you or anyone you know be interested in an enhancer trap mouse that has expression in a discrete region of each limb as well as the face? I hesitate to kill these mice off because I can't help but think someone, somewhere might be interested.

(2) Do you know of a bulletin board for posting pictures of enhancer trap (gene trap etc) mice? I think such a board would be quite valuable. For example, we have genrated several enhancer trap mice (one that stained a region of the brain and the spinal cord is regrettably gone). None of these mice are of interest to us since we are looking for a SMC-restricted pattern of expression (see SM22 transgenic mouse paper in March '96 issue of JCB). There may be others, however, who would gladly accept such mice.

Sincerely,

Joseph M. Miano, Ph.D.

From kelly at citi2.fr Thu Oct 17 21:42:58 1996 From: kelly at citi2.fr (U344) Date: Wed Aug 20 11:52:53 2003 Subject: knockout mouse construct problems Message-ID:

> At this stage we are working with a very large piece of DNA >(~12KB)- and perhaps this will lower the ligation frequency-but we have >generated larger plasmids with little problem. > We are using JM109 or DH5a as our competent cells (chemically >competent). Any suggestions at all would be greatly appreciated!

Whenever I'm forced to deal with DNA of such a size, I use the Hanahan transformation proceedure to produce competent cells... the original reference is J. Mol. Biol. 1983, (166) pg 557... it may also be to your advantage to lift and screen colonies before picking them.

good luck...

INSERM Unite 344 Endocrinologie Moleculaire 156 rue de Vaugirard, 75730 Paris cedex 15 France 'Les raisonables ont dure, les passiones ont vecu'

From ldeg at midway.uchicago.edu Fri Oct 18 18:19:37 1996 From: ldeg at midway.uchicago.edu (Linda Degenstein) Date: Wed Aug 20 11:52:53 2003 Subject: Transgenic Facility Yellow Pages Message-ID: <[email protected]>

I am often contacted by people outside my university inquiring as to whether or not we can produce KO or transgenic mice for them. These are often from investigators out of the state or even out of the country. I try to match them up with an institution in their location if I know of one.

I would like to make a list of all the known facilities, with pertinent info, such as institution, location, services offered, contact person, telephone and e-mail listings, and whether or not you can perform these services for someone outside your institution, but in your area.

If you are willing to e-mail this info, I will summarize in about a week for both lists.

Thank you for your help.

From pinkert at cmed.bhs.uab.edu Fri Oct 18 18:31:33 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert) Date: Wed Aug 20 11:52:53 2003 Subject: Transgenic Facility Yellow Pages Message-ID: <[email protected]>

> Date: Fri, 18 Oct 1996 17:19:37 -0500 (CDT) > To: [email protected], [email protected] > From: [email protected] (Linda Degenstein) > Subject: Transgenic Facility Yellow Pages > Reply-to: [email protected] > I am often contacted by people outside my university inquiring as to > whether or not we can produce KO or transgenic mice for them. These are > often from investigators out of the state or even out of the country. I try > to match them up with an institution in their location if I know of one. > > I would like to make a list of all the known facilities, with pertinent > info, such as institution, location, services offered, contact person, > telephone and e-mail listings I think you have info for the NICHD facility at UAB. I'm not sure if such a listing of all facilities will be problematic with the patent positions that DNX (as well as GenPharm, Lexicon, etc. for ES cell work) are staking out. But so long as the caution is there in advance, it may all be OK.

From a.annala at ucl.ac.uk Sun Oct 20 14:32:43 1996 From: a.annala at ucl.ac.uk (Dr. Alexander J. Annala) Date: Wed Aug 20 11:52:53 2003 Subject: Transgenic Facility Yellow Pages Message-ID:

Dr. Pinkert,

I am curious about the patent claims you mention. We do some pronuclear microinjection of DNA, ES cell injection into blastocysts, and aggregation of ES cells with morula to produce transgenic mice. Are individuals and/or organizations claiming patent rights which would preclude carrying out these basic transgenic procedures in academic laboratories? Are license fees to obtain the right to perform these procedures anywhere within the reach of small academic laboratories?

Are the basic transgenic patent claims analogous to the original Stanford recombinant DNA claims where a $10,000 fee was required for any lab to perform basic molecular biology techniques? It would seem few, if any, academic laboratories pay this fee today.

It might be interesting for people to have an up to date listing available on the network containing a summary of transgenic relevant patent claims and licensing arrangements/fees.

Thanks,

Alexander J. Annala, Ph.D. Wellcome Senior Research Fellow Laboratory for Molecular Pharmacology University College London Gower Street London WC1E 6BT

Tel: +44(171)380-7857 Fax: +44(171)380-7245 Email: [email protected]

>> Date: Fri, 18 Oct 1996 17:19:37 -0500 (CDT) >> To: [email protected], [email protected] >> From: [email protected] (Linda Degenstein) >> Subject: Transgenic Facility Yellow Pages >> Reply-to: [email protected] > >> I am often contacted by people outside my university inquiring as to >> whether or not we can produce KO or transgenic mice for them. These are >> often from investigators out of the state or even out of the country. I try >> to match them up with an institution in their location if I know of one. >> >> I would like to make a list of all the known facilities, with pertinent >> info, such as institution, location, services offered, contact person, >> telephone and e-mail listings >I think you have info for the NICHD facility at UAB. I'm not sure if >such a listing of all facilities will be problematic with the patent >positions that DNX >(as well as GenPharm, Lexicon, etc. for ES cell work) are staking >out. But so long as the caution is there in advance, it may all be OK. > >Carl

From pinkert at cmed.bhs.uab.edu Sun Oct 20 10:02:00 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert) Date: Wed Aug 20 11:52:53 2003 Subject: Transgenic Facility Yellow Pages Message-ID: <[email protected]>

> Dr. Pinkert, > > I am curious about the patent claims you mention. We do some pronuclear > microinjection of DNA, ES cell injection into blastocysts, and aggregation > of ES cells with morula to produce transgenic mice. Are individuals and/or > organizations claiming patent rights which would preclude carrying out > these basic transgenic procedures in academic laboratories? If you are providing services in house, there should be no problem. If you provide services outside your home institution, you can be liable to the folks holding patents on all the microinjection/gene transfer patents that have been awarded. Are license > fees to obtain the right to perform these procedures anywhere within the > reach of small academic laboratories? When it is allowed by the patent holders. But if the work is in-house AND for non-commercial basic research - most of the companies with the rights will leave you alone. It is only when you offer a service or begin toward commercialization that you will find yourself in need of appropriate legal counsel. > > Are the basic transgenic patent claims analogous to the original Stanford > recombinant DNA claims where a $10,000 fee was required for any lab > to perform basic molecular biology techniques? It would seem few, if > any, academic laboratories pay this fee today. Agreed, but as an example, for DNA microinjection outside of your home institution, DNX would not allow you to provide the service (as it competed with their program - but in-house basic research was automatically exempted). Now, if you want to commercialize or transfer animals, they have a basic fee structure in mind. However, if you agree to provide microinjection services to say another institution - even at/below your actual costs, you would likely be liable for treble damages . . . > > It might be interesting for people to have an up to date listing available on > the network containing a summary of transgenic relevant patent claims > and licensing arrangements/fees. I agree, but some of the ES cell patents as well as more recent targeting/expression patents are in process and disclosure is not readily available. > Good luck.

Carl A. Pinkert

From stewarv at cesmtp.ccf.org Mon Oct 21 12:27:16 1996 From: stewarv at cesmtp.ccf.org (Valerie Stewart MS)

Subject: Transgenic Facility Yellow Pages -Reply Message-ID:

Hi Linda--

Good idea, though with the patent battles, our reach is limited. Our policy so far has been to make mice for Clinic investigators and their collaborators. Here is the information you requested:

Transgenic/IKnockout Core Facility Cleveland Clinic Research Institute FF6-50, 9500 Euclid Avenue Cleveland, OH 44106 Director, Valerie Stewart Technician, Christine Brant phone (216)444-8521 fax (216)445-6257 e-mail "[email protected]" Services include pronuclear injection, blastocyst injection, cryopreservation by vitrification, training in all techniques. We also do microinjection of proteins, etc. into cell lines via an Eppendorf automated system. Services and prices can also be accessed through the Clinic's web page at "www.ccf.org". Go to Main Menu, then Research Institute, then Scientific Support Services, then Core Services.

>>> Linda Degenstein - 10/18/96 6:19 PM >>> I am often contacted by people outside my university inquiring as to whether or not we can produce KO or transgenic mice for them. These are often from investigators out of the state or even out of the country. I try to match them up with an institution in their location if I know of one.

From YOCKEY at DCSMSERVER.MED.SC.EDU Mon Oct 21 19:15:02 1996 From: YOCKEY at DCSMSERVER.MED.SC.EDU (Courtland E. Yockey)

Subject: selection cassette-dependent phenotypes Message-ID: <[email protected]>

Hello,

I have a question regarding the phenotypes of targeted mutant mice.

One justification for flanking selection/mutation cassettes with loxP sites is so that the cassette - and the transcription unit associated with the selectable marker - can be removed in order to avoid an artifactual mutant phenotype that is dependent upon its presence.

Does anyone know of published cases in which the phenotype of a mutant mouse is influenced by the presence of the selectable marker itself within the mutant locus? Cases in which the phenotype was compared, for instance, before and after Cre-mediated excision of a loxP-flanked cassette?

Thanks for the information.

Courtland Yockey ------Courtland Yockey, M.S. from the lab of Noriko Shimizu, Ph.D.

University of South Carolina School of Medicine Department of Developmental Biology & Anatomy Columbia, SC 29208 USA

Phone# 803-733-1503 Fax# 803-733-1533

From dolle at titus.u-strasbg.fr Tue Oct 22 15:19:24 1996 From: dolle at titus.u-strasbg.fr (Pascal DOLLE)

Subject: selection cassette-dependent phenotypes Message-ID: <[email protected] >

>Hello, > >I have a question regarding the phenotypes of targeted mutant mice. > >One justification for flanking selection/mutation cassettes with loxP >sites is so that the cassette - and the transcription unit associated >with the selectable marker - can be removed in order to avoid an >artifactual mutant phenotype that is dependent upon its presence. > >Does anyone know of published cases in which the phenotype of a mutant >mouse is influenced by the presence of the selectable marker itself >within the mutant locus? Cases in which the phenotype was compared, for >instance, before and after Cre-mediated excision of a loxP-flanked >cassette? > >Thanks for the information. > >Courtland Yockey >------

Dear Courtland, a couple of years ago, we have reported that some 'weird' & dominant phenotypes in chimeras harboring a Hoxd-10 disruption may have resulted from integration effect of the neo cassette in the locus. At that time, the loxP technique was not developed, so we didn't have this mutation without the selection cassette. The ref is Rijli et al., Dev. Dynamics 201, p. 366 (1994).

I guess there will be more and more cases of mutations with or without neo cassettes coming out in the next future. You could contact Filippo Rijli in our institute who may have other examples of lines with or without TK- neo integrations.

Sincerely,

Pascal DOLLE, M.D., Ph.D.

CNRS, Unite 184 INSERM I.G.B.M.C. Parc d'innovation B.P. 163 67404 ILLKIRCH Cedex FRANCE

Tel: (33) 88 65 33 34 or 40 Fax: (33) 88 65 32 01

From dbowtell at petermac.unimelb.edu.au Wed Oct 23 09:50:38 1996 From: dbowtell at petermac.unimelb.edu.au (David Bowtell)

Subject: selection cassette-dependent phenotypes In-Reply-To: <[email protected] > Message-ID:

As a follow on to the discussion about selectable markers we have placed a lacZneo fusion in frame downstream of the Sos1 gene. The marker is driven by the Sos1 promoter and there is no heterologous promoter sequences inserted. We appear to see, however, mosaic expression of lacZ in some heterozygous animals and wondered about silencing of the targeted locus in some tissues. Has anyone else encountered this?

From anna at screams.gdb.org Wed Oct 23 10:29:45 1996 From: anna at screams.gdb.org (Anna Anagnostopoulos)

Subject: Mycoplasma Infection Message-ID:

Hello everyone,

Dr. Yasuzawa is trying to rid his mice of a Mycoplasma infection. I am sure he would appreciate additional suggestions.

Thanks a lot

Anna V. Anagnostopoulos

> The reason why I noticed is that my mice are infected by mycoplasma. > Especially, the clinical signs of old mice are more serious than young. > I'm worrying about both sudden death and expansion of contamination. > I already start to feed tetracyclin to them. But, the condition has not > improved yet. > If you give me any suggestion how to rescue them, I'm very happy. > > > yours sincerely, > > Kayoko Yasuzawa > > > > >

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ Anna V. Anagnostopoulos, Ph.D. Division of Biomedical Information Sciences The Johns Hopkins University School of Medicine 2024 E. Monument Street Baltimore, MD 21205-2236 tel: (410) 614-3226 fax: (410) 614-0434 e-mail: [email protected] From milstone at rascal.med.harvard.edu Wed Oct 23 12:26:12 1996 From: milstone at rascal.med.harvard.edu (David Milstone)

Subject: selection cassette-dependent phenotypes Message-ID:

>Does anyone know of published cases in which the phenotype of a mutant >mouse is influenced by the presence of the selectable marker itself >within the mutant locus? Cases in which the phenotype was compared, for >instance, before and after Cre-mediated excision of a loxP-flanked >cassette? >> >Courtland Yockey

Selectable markers have been shown to influence the phenotype of mutant mice by cis-mediated effects on transciptional regulation (published examples include beta globin LCR element and CD11b). But this is presumably distinct from effects of the coding region sequences of the selectable marker itself.

David S. Milstone Vascular Research Division Department of Pathology Brigham & Women's Hospital LMRC 421 221 Longwood Avenue Boston, MA USA

From r-carver at nimr.mrc.ac.uk Thu Oct 24 17:40:21 1996 From: r-carver at nimr.mrc.ac.uk (Rick Carver)

Subject: Mycoplasma Infection Message-ID: <[email protected]>

Dear All

>Dr. Yasuzawa is trying to rid his mice of a Mycoplasma infection. I am >sure he would appreciate additional suggestions. > >> The reason why I noticed is that my mice are infected by mycoplasma. >> Especially, the clinical signs of old mice are more serious than young. >> I'm worrying about both sudden death and expansion of contamination. >> I already start to feed tetracyclin to them. But, the condition has not >> improved yet.

My advice would be to rederive them, preferably by embryo transfer. Antibiosis is a poor option because you can never be certain that it has been fully effective and you run a major risk of producing tetracycline-resistant Mycoplasma. The NRC's 'Infectious Diseases of Mice and Rats' has a nice description of the protocol to be followed post-caesarean rederivation. This protocol is equally valid if you rederive by embryo transfer. ET is much more microbiologically secure and if he is doing transgenic work he should already have the necessary equipment and skills available.

I hope that this is of assistance.

From dknudsen at scruznet.com Wed Oct 23 23:16:18 1996 From: dknudsen at scruznet.com (Dave Knudsen)

>Hello everyone, > >Dr. Yasuzawa is trying to rid his mice of a Mycoplasma infection. I am >sure he would appreciate additional suggestions. > >Thanks a lot > >Anna V. Anagnostopoulos > >> The reason why I noticed is that my mice are infected by mycoplasma. >> Especially, the clinical signs of old mice are more serious than young. >> I'm worrying about both sudden death and expansion of contamination. >> I already start to feed tetracyclin to them. But, the condition has not >> improved yet. >> If you give me any suggestion how to rescue them, I'm very happy.

Anna -

I would agree with Rick Carver in that rederivation is the best way to proceed for a contaminated colony. The assumption under this agreement is that the affected colony is a small breeding colony, and not a group of stock animals that could be eradicated and replaced at the end of an experiment. I also assume that your colleague, Dr. Yasuzawa, has completed the diagnostic process and confirmed mycoplasmosis as the problem. This diagnosis can be made by serology, culture, or histopathology (preferably all three for a colony, to reduce the chance of false negative findings). The confirmation itself is important to resolution of the problem - I can think of several instances where other opportunistic pathogens, environmental factors, immunodeficiencies, and/or transgenic phenotypes have mimicked the clinical presentation of murine mycoplasmosis quite closely. Also, I would add that antibiosis may help the clinical appearance of the animals by temporarily arresting disease progression, but it will do nothing in evoking a cure, and treated animals will continue to be carriers to infect new introductions (and births) into the colony.

I would personally be most comfortable, however, in dealing with the rederivation issue by caesarian section; I suspect that confidence is gained from the method most familiar. Over the years I have had more positive experiences with caesarian rederivation in eliminating pathogens from mouse colonies, and embryo or ovarian manipulations seem always to break at some point. Perhaps the best method could be determined by close questioning of the veterinary staff of the affected institution and working with them to resolve the outbreak.

Dave Knudsen DVM, DACLAM Scotts Valley CA, USA

From pasceri at sickkids.on.ca Tue Oct 1 21:34:58 1996 From: pasceri at sickkids.on.ca (Peter Pasceri)

Subject: Cryopreservation Message-ID:

Hi all! I am looking for suppliers of mouse embryo freezing kits. Can you help?

Thanks,

-- Peter Pasceri [email protected] Genetics Research Phone: (416) 813-4205 The Hospital for Sick Children Toronto, Ontario,Canada

From p.sobieszczuk at ic.ac.uk Thu Oct 3 11:37:29 1996 From: p.sobieszczuk at ic.ac.uk (Peter Sobieszczuk)

Subject: Transgenic mice/Transplantation (fwd) Message-ID:

Sorry, if it is a duplication for some of you. Peter (your listowner)

>Hello everyone, > >Any advise/suggestions on the following questions would be greatly >appreciated. > >Thank you > >Anna V. Anagnostopoulos >TBASE >Division of Biomedical Information Sciences >The Johns Hopkins University School of Medicine >[email protected] > >------> >I am working with mice, doing Bone Marrow and Peripheral Blood >Tranplantation experiments, at the Department of Immunogenetics of the New >York Blood Center. We are interested in developing a syngeneic mouse >transplantation model. We would like to use as donors, animals carrying a >transgene, that its product is not lethal or significantly affecting the >recipients (especially hematologically), and also is easily detectable >post-transplant. For example, mice transgenic for a human gene, encoding >for a human antibody, would be a good model. The idea would be to show >that the donor cells (positive for the transgene) engraft, and the >transgene is functional (we can detect the product) after transplant. The >mouse strains I am working with at the present time are C57BL/6, A/J and >B6A (C57BL/6 X A/J) (all obtained from Jackson Labs). It would help >tremendously if the transgenic animals are of the same background. Do you >have any suggestions? >

From mangia at uniroma1.it Fri Oct 4 12:46:36 1996 From: mangia at uniroma1.it (Franco Mangia)

Subject: Transgenic mice/Transplantation (fwd) Message-ID:

> >We are interested in developing a syngeneic mouse >>transplantation model. We would like to use as donors, animals carrying a >>transgene, that its product is not lethal or significantly affecting the >>recipients (especially hematologically), and also is easily detectable >>post-transplant. For example, mice transgenic for a human gene, encoding >>for a human antibody, would be a good model. The idea would be to show >>that the donor cells (positive for the transgene) engraft, and the >>transgene is functional (we can detect the product) after transplant. The >>mouse strains I am working with at the present time are C57BL/6, A/J and >>B6A (C57BL/6 X A/J) (all obtained from Jackson Labs). It would help >>tremendously if the transgenic animals are of the same background. Do you >>have any suggestions? >> >Anna V. Anagnostopoulos >TBASE >Division of Biomedical Information Sciences >The Johns Hopkins University School of Medicine >[email protected]

I think that the ROSA26 transgenic mice, developed by Phil Soriano, are just what you need for labeling engrafts at a single cell level. These mice carry a lacZ transgene directed by a ubiquitous promoter active in all tissues, without apparently affecting cell viability, since these mice are fine. These mice can be obtained from the Jackson Labs either as B6,129 F1 (the original Soriano's hybrid strain) or with a pure C57BL/6J background. Complete names of these mice are: "B6,129-TgR(ROSA26)26 Sor", and "C57Bl/6J-TgR(ROSA26)26 Sor". I am also very interested in obtaining an ES cell line(s) from these mice. Is anybody aware whether these ES lines are being developed in some lab? Also, does anybody know whether ROSA26 mice with nuclearly targeted beta-gal have been produced? Franco

Franco Mangia Laboratory of General Biology Department of Psychology University La Sapienza of Rome c/o Institute of Histology and General Embryology Via Borelli, 50 00161 Rome, Italy tel: 39-6-4976-8103 FAX: 39-6-4976-8099 E-mail: [email protected]

From p.sobieszczuk at ic.ac.uk Sat Oct 5 18:19:10 1996 From: p.sobieszczuk at ic.ac.uk (Peter Sobieszczuk)

Subject: "TgList Backstage" (admin. news *2) ARCHIVES & TBASE Message-ID:

Dear Transgenic Subscribers,

Greetings to newcomers,

ARCHIVES Charmaine Foltz asked me some time ago: >I was wondering if the information on the list will be retrievable >(i.e., will there be a data base that we can search archived files). very much so, send the message: index transgenic-list to listserver at [email protected] and you shall receive an index of monthly files, something like:

>>>> index transgenic-list total 241 -rw-rw---- 1 daemon 10364 Jul 29 16:38 transgenic- list.archive.9607 -rw-rw---- 1 majordom 146666 Aug 29 21:36 transgenic- list.archive.9608 -rw-rw---- 1 majordom 71873 Sep 25 15:31 transgenic- list.archive.9609 -rw-rw---- 1 majordom 6715 Oct 4 12:46 transgenic- list.archive.9610 >>>> where July entries file name is "transgenic-list.archive.9607"

If you then send: get transgenic-list transgenic-list.archive.9607 you should receive all postings for that month (in this case, July) as individual e-mail messages.

If you have access to world wide web, (WWW) you can explore archives with the luxury of selecting individual entries etc., using your favourite browser, Netscape, Mosaic or Explorer.

The URL address is: http://www.lists.ic.ac.uk/hypermail/transgenic-list

I'm afraid however that even with the browser, files are not searchable (at this stage anyway), although subject, (which should be the topic of any message) is clearly visible.

TBASE Anna Anagnostopoulos who is co-ordinating TBASE data base site sent this introduction (with minor editorial changes P.S.):

------BEGINNING OF TEXT

TBASE, the Transgenic/Targeted Mutation Database, is an attempt to organize information on transgenic animals and targeted mutations generated, and analyzed worldwide. TBASE is available on the WWW http://www.gdb.org/Dan/tbase/tbase.html and can be searched either quite broadly across the entire database (the 'blunt object' approach) or quite specifically with the use of 28 distinct searchable fields (the 'scalpel and tweezers' approach). TBASE contains a wealth of information about each animal in the database. A selection of the fields present in the databases are: line name, genus, DNA construct description, host background, line source, method used , transgene/targeted allele expression, lethality, phenotype, handling, contact information, full citation and abstracts of the papers related to the creation of a line.

Owing to the wide use of homologous recombination in ES cells, TBASE initially focused on maintaining a comprehensive coverage of all targeted mutations reported to date. Moreover, it focused on the mouse as the predominant mammalian model. Specific emphasis is still placed on cataloging novel knockout mice, which have defined genetic modifications, with an extensive detailed description of their resulting mutant phenotypes. In the future, the focus of TBASE will expand to cover all animals resulting from transgenesis or targeted mutation, regardless of species. Additionally, TBASE plans to incorporate 'transgenic knockouts', that is, knockout animals that have subsequently served as recipients for transgenes.

TBASE should be viewed as a dynamic database with a potentially unlimited number of entries, expected to grow considerably as more experimental methods and lines are generated. Data acquisition is primarily effected by active literature scanning and manual data entry, as well as processing of direct submission forms.

Literature scanning refers to direct data extraction from the scientific literature through regular examination of over 20 journals. These journals have been statistically identified as periodically stable sources of information on transgenesis and gene targeting. Active literature scanning has, so far, been the primary mode of data accumulation, and has ensured that the database faithfully reflects the general direction of related research. It is expected to remain a significant component of the data acquisition tactics, even as other methods of data acquisition become available (see below). In an effort to avoid missing important data, keyword-related references from additional journals are also screened by the use of Medline and Current Contents.

In the future, data that are not manually entered by TBASE staff through the literature scanning process, are expected to arrive either as direct paper or as electronic submissions. Although over 98% of the data have been entered manually so far, direct paper submissions are expected to increase as TBASE becomes fully adopted by the scientific community. In addition, as electronic submission tools become available, more research groups will be prepared to commit resources to the production of data deposited into TBASE. Electronic submission tools and their error-checking capabilities will ensure efficient and timely deposition of the data.

END OF TEXT ------

STATISTIC: There are approx. 240 members from at least 18 countries registered now, and I'm starting to think aren't we growing too big, read to much to read. Regards,

Yours sincerely,

Listowner

______

Dr Peter Sobieszczuk Imperial College School of Medicine at St. Mary's tel: 0171 594 3784 Biochemistry and Molecular Genetics tel: 0171 594 3799 Norfolk Place fax: 0171 706 3272 London W2 1PG, U.K. e-mail: [email protected] ______

From khelmuth at facstaff.wisc.edu Wed Oct 9 15:28:47 1996 From: khelmuth at facstaff.wisc.edu ([email protected])

Subject: fire codes within animal colony Message-ID:

Hello everyone-

From landel at medsci.udel.edu Wed Oct 9 15:44:03 1996 From: landel at medsci.udel.edu (Carlisle Landel)

Subject: fire codes within animal colony Message-ID: <[email protected]>

I'm curious--how often do these alarms go off? Maybe instead you could get them to absolve you from frequent fire drills.

Carlisle Landel

>From [email protected] Wed Oct 9 10:35:44 1996 >Mime-Version: 1.0 >Content-Type: text/plain; charset="us-ascii" >To: [email protected] >From: [email protected] >Subject: fire codes within animal colony >Sender: [email protected] >Precedence: bulk >Reply-To: [email protected] > >Hello everyone- > >I face a delemma within our facility and am hoping someone can help me out. >We recently moved our rat and mouse colonies into a new building. >Everything seems to be going well except for one thing. We experienced the >first fire drill and realized that there are 3 fire alarms within our 5ft. >wide clean hallway and 3 fire alarms within our 5ft. wide dirty hallway. >They are extremely loud and potentially problematic for our animals. Our >building manager agrees that this seems extremely overdone and wishes to >get some of the alarms diconnected. However, the state fire code regulator >feels that with the background noises of the cage washer, movement of carts >and racks, etc. that these extra fire alarms are necessary. He will only >follow suit if I can come up with evidence that not only does it affect the >well-being of the animals but that other facilities have a small number of >alarms. I have obtained documentation of loud noises and it's effect on >breeding, embryo production, etc. Can someone help me out with regulations >regarding fire alarms within your animal facility? > >Any suggestion on how to deal with this would be beneficial. >Thanks > >Kathy Krentz Helmuth > >Kathy Krentz Helmuth >Transgenic Facility >425 Henry Mall >Room 2210 Biotechnology Center >Madison, WI 53706 >(608) 265-2801 >email:[email protected] > > > > >

From duffyh at war.wyeth.com Wed Oct 9 19:45:06 1996 From: duffyh at war.wyeth.com (Heidi Duffy)

Subject: fire codes within animal colony -Reply Message-ID:

I used to work in animal facility that used flashing lights instead of audible alarms within the animal facility. The audible alarms were only in the office areas of the facility. (Our phones within the facility were set up the same way, including the cagewash areas). Designated techs were assigned to check all animal rooms for personnel evacuation before leaving the buildings. It worked out very well.

Heidi Duffy email:[email protected] From david_gask at Merck.Com Thu Oct 10 13:47:00 1996 From: david_gask at Merck.Com (David Gask)

Subject: Fire Alarm problems-Kathy Krentz Helmuth Message-ID: <199610101150.HAA21899@igw2>

Kathy, Why don't you investigate the use of ' SILENTONE ALARMS' as an alternative. These are used widely in the UK and were developed to prevent problems for the animals caused by more conventional sounders. They produce a loud noise suitable for use as an alarm in the event of fire, but at the same time the frequency is below the threshold frequency, or outside the most sensitive range for most commonly used laboratory animals. If you require the address of a supplier I can send it to you. All our animal facility fire alarms are of this type.

Hope this helps

Dave Gask Operations Coordinator Merck Sharp & Dohme Neuroscience Research Centre Terlings Park Eastwick Road Harlow, Essex CM20 2QR England Tel 01279 440220 e-mail [email protected]

Hello everyone-

From Roger.Leemann at imr.psi.ch Thu Oct 10 14:21:53 1996 From: Roger.Leemann at imr.psi.ch (Roger C. Leemann)

Subject: Rodent-research list Message-ID:

Since I'm working with mice but do not use transgenic animals, I subscribed to the rodent-research list about a month ago. Majordomo told me, that it had forwarded my request to the list owner for approval but I haven't got any response since. Maybe it's not a list for every Jack and Joe (or was it Jack and Jill?), or maybe the list owner is too busy, or on vacation, whatever. I don't mind if they don't want me, but ones likes to know... Anybody in this list who has a clue why the rodent-research list seems dead?

::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::: ::::: Work: Institute for Medical Radiobiology ++41 (56) 310-3792 [desk] CH-5232 Villigen PSI ++41 (1) 3856-561 [desk ZH] Switzerland ++41 (56) 310 3294 [fax] Home: Nordstrasse 26, CH-8006 Zuerich Switzerland ++41 (1) 361 0349 [home] From p.sobieszczuk at ic.ac.uk Thu Oct 10 16:55:30 1996 From: p.sobieszczuk at ic.ac.uk (Peter Sobieszczuk)

Subject: Rodent-research list & "TgL Backstage"(admin.news*3) Message-ID:

Dear Roger,

>Since I'm working with mice but do not use transgenic animals, I subscribed >to the rodent-research list about a month ago. Majordomo told me, that it I presume listserver "majordomo" at Caltech, and not "ic.ac.uk" at Imperial College?

>Maybe it's not a list for every Jack and Joe (or was it Jack and Jill?), or Do not take it personnally, it's just a computer.

>maybe the list owner is too busy, or on vacation, whatever. I don't mind if >they don't want me, but ones likes to know...

Sorry, I still do not know what exactly happened, Eric Mercer had some technical problems supporting "rodent-research" list, 3 or 4 months ago. A few people has tried to contact him, at the end, this list was created (see the introduction to the list available also by sending "info transgenic-list" message to "[email protected]" address).

You have been with us (transgenic-list) for the month now and probably noticed that the list is un-moderated and has no restriction on the content. Transgenic - is just a name on a server, and the list a quick way to exchange information.

I would not like to start a discussion on definition of transgenesis, but would imagine that for the active researchers in the field it would cover quite a bit of biology, from molecular to developmental, with the animal models not exclussively restricted to rodents.

So it is not the name but participants who will take the discussion in any particular direction, including yourself.

Who would you like to see on the list Roger, or should I ask everybody:

Who else do you think we should invite to the list?

Best regards,

Sincerely,

Peter - your listowner

From s.tan at anatomy.unimelb.edu.au Tue Oct 15 05:09:53 1996 From: s.tan at anatomy.unimelb.edu.au (S.Tan)

Subject: Fostering baby mice Message-ID:

I am grafting pieces of neural tissue into the cortex of newborn mice but find that the mother keeps eating the babies. The wound is closed with a single silk suture and this appears to attract the attention of mum. The mother's strain is C57/BL6 x DBA. Are there less aggressive strains of mums out there? Thanks.

From a.annala at ucl.ac.uk Tue Oct 15 06:51:24 1996 From: a.annala at ucl.ac.uk (Dr. Alexander J. Annala)

Subject: Fostering baby mice Message-ID:

>I am grafting pieces of neural tissue into the cortex of newborn mice but >find that the mother keeps eating the babies. The wound is closed with a >single silk suture and this appears to attract the attention of mum. The >mother's strain is C57/BL6 x DBA. Are there less aggressive strains of >mums out there? Thanks.

Most F1's make very good mothers. Your C57BL/6 x DBA are F1 mice. We use C57BL/6 x CBA and C57BL/10 x CBA with good results.

Do you take precaution to avoid leaving scent on operated baby mice? Some people roll babies in mom's urine before returning them to the nest. Others treat mom's nose with alcohol to keep her from smelling any differences between babies for awhile. This might be a scent issue rather than an exposed suture problem.

Internalize suture might be difficult in baby mice. Maybe you could make your incision somewhere away from your skull opening, slide the skin opening over the area you want to open bone, do your work, slide the skin back, and maybe use one of the tissue adhesives which are pretty available around medical schools and operating theatres to close the wound. This would avoid leaving exposed suture for the mom to chew on. From fobo at ltk.unizh.ch Tue Oct 15 07:22:40 1996 From: fobo at ltk.unizh.ch (Frank Bootz)

Subject: Fostering baby mice Message-ID: <[email protected]>

Dear colleaque,

I have done thymectomy in newborn mouse. The wound was also closed with a single U-suture in the neck. The pups awake within 5 minutes after methofane (methoxyflurane) anesthesia and brought back to the mother immediately. We don't anesthetize the mother, contrary the most papers describing this procedure. In this case the mother's strain is C57/Bl/6 x PO. If we can choose the strain, we normally use Zur:ICR original Charles River swiss mice CD-1. They are mice with excellent mother attributes.

Best regards,

Frank Frank Bootz Institute of Laboratory Animal Science University of Zurich

Winterthurerstrasse 190 phone: +41 1 257 54 55 8057 Zurich fax: +41 1 257 57 03 Switzerland e-mail [email protected]

From GWOLFF at NCTR.FDA.GOV Tue Oct 15 14:44:51 1996 From: GWOLFF at NCTR.FDA.GOV (George L. Wolff)

Subject: Fostering baby mice Message-ID: <[email protected]>

Zur:ICR cannot be descended from CD-1 mice - or the strain designation is incorrect. Let me know if you'd like more details. George L. Wolff

Tel: (501) 543-7522 FAX: (501) 543-7635/7662 NCTR HFT-140 3900 NCTR Road Jefferson, AR 72079-9502 From gotto at leland.Stanford.EDU Wed Oct 16 20:03:00 1996 From: gotto at leland.Stanford.EDU (Glen Otto)

Subject: Fostering baby mice In-Reply-To: <[email protected]> Message-ID:

A non-text attachment was scrubbed... Name: not available Type: text/enriched Size: 577 bytes Desc: not available Url : http://mailman.ic.ac.uk/mailman/private/transgenic- list/attachments/19961016/3cf223e6/attachment-0001.bin From a.annala at ucl.ac.uk Thu Oct 17 05:50:59 1996 From: a.annala at ucl.ac.uk (Dr. Alexander J. Annala)

Subject: Fostering baby mice -- Failure to thrive? Message-ID:

What can be done to assist baby mice in distress after birth? Some chimeras with neuroreceptor channel mutations experience difficulty feeding themselves -- and sometimes even suckling milk from mother.

We have in the past injected approx 20% weight of animal with Ringers Lactate (Hartman's Solution). However there must be more aggressive therapy which would facilitate survival of distressed baby mice.

We have also used incubator -- with reservations about the possibility of dehydration from elevated environmental temperature.

Thanks,

Alexander J. Annala, Ph.D. Senior Research Fellow Laboratory for Molecular Pharmacology University College London

From carton at murray.fordham.edu Wed Oct 16 23:44:40 1996 From: carton at murray.fordham.edu (Jill Carton)

Subject: knockout mouse construct problems Message-ID:

We are working on generating several knockout mice and have been successful in producing transgenes for some of our genes, but we have a fairly consistent problem with the final ligation step of assembling the construct. When we PCR our ligation reaction to see if the DNA pieces have ligated we do get the appropriate product however analysis of the clones by miniprep show no positive clones. So we are suspecting that the problem is in the uptake or growth of the construct in the bacterial cells. Although, each fragment of the transgene can be propagated in the bacteria. At this stage we are working with a very large piece of DNA (~12KB)- and perhaps this will lower the ligation frequency-but we have generated larger plasmids with little problem. We are using JM109 or DH5a as our competent cells (chemically competent). Any suggestions at all would be greatly appreciated!

From p.sobieszczuk at ic.ac.uk Thu Oct 17 15:23:53 1996 From: p.sobieszczuk at ic.ac.uk (Peter Sobieszczuk)

Subject: Nod/Scid mice Message-ID:

Forwarded from [Anna Anagnostopoulos ]

>Hello everyone, > >Dr. Sally J. Cutler (Department of Medical Microbiology, Charing Cross >Hospital, London. Telephone:+ 44181 846 7570) is looking for sources of >Nod/Scid mice, preferably in the UK. Any suggestions on where to look? > >Thank so much > >~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ >Anna V. Anagnostopoulos, Ph.D. >Division of Biomedical Information Sciences >The Johns Hopkins University School of Medicine >2024 E. Monument Street >Baltimore, MD 21205-2236 >tel: (410) 614-3226 >fax: (410) 614-0434 >e-mail: [email protected]

From james at icr.ac.uk Thu Oct 17 22:00:12 1996 From: james at icr.ac.uk (james wallace) Subject: Nod/Scid mice Message-ID:

> >Dr. Sally J. Cutler (Department of Medical Microbiology, Charing Cross > >Hospital, London. Telephone:+ 44181 846 7570) is looking for sources of > >Nod/Scid mice, preferably in the UK. Any suggestions on where to look? > > > >Thank so much > > > >~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ > >Anna V. Anagnostopoulos, Ph.D. > >Division of Biomedical Information Sciences > >The Johns Hopkins University School of Medicine > >2024 E. Monument Street > >Baltimore, MD 21205-2236 > >tel: (410) 614-3226 > >fax: (410) 614-0434 > >e-mail: [email protected] > > Hi, Tell Sally to give me a call as we have a colony of Nod\Scids here at ICR in the UK.

Best Wishes

Jim Wallace Institute of Cancer Research The McElwain Laboratories 15 Cotswold Rd Belmont Sutton Surrey SM2 5NG England tel: 0181 643 8901 Ext 4638 Fax: 0181 770 1395 e_mail [email protected] > >

From anna at screams.gdb.org Thu Oct 17 18:18:07 1996 From: anna at screams.gdb.org (Anna Anagnostopoulos)

Subject: Available enhancer traps Message-ID: Hello again,

If any of you is interested in Joe Miano's enhancer traps or knows of a bulletin where he can have the pictures posted please let me know.

------Forwarded message ------Date: Wed, 16 Oct 1996 16:36:44 -0500 From: Joe Miano To: [email protected] Subject: enhancer trap

We do transgenics and I have two questions:

(1) Would you or anyone you know be interested in an enhancer trap mouse that has expression in a discrete region of each limb as well as the face? I hesitate to kill these mice off because I can't help but think someone, somewhere might be interested.

(2) Do you know of a bulletin board for posting pictures of enhancer trap (gene trap etc) mice? I think such a board would be quite valuable. For example, we have genrated several enhancer trap mice (one that stained a region of the brain and the spinal cord is regrettably gone). None of these mice are of interest to us since we are looking for a SMC-restricted pattern of expression (see SM22 transgenic mouse paper in March '96 issue of JCB). There may be others, however, who would gladly accept such mice.

Sincerely,

Joseph M. Miano, Ph.D.

From kelly at citi2.fr Fri Oct 18 05:42:58 1996 From: kelly at citi2.fr (U344)

Subject: knockout mouse construct problems Message-ID:

> At this stage we are working with a very large piece of DNA >(~12KB)- and perhaps this will lower the ligation frequency-but we have >generated larger plasmids with little problem. > We are using JM109 or DH5a as our competent cells (chemically >competent). Any suggestions at all would be greatly appreciated!

Whenever I'm forced to deal with DNA of such a size, I use the Hanahan transformation proceedure to produce competent cells... the original reference is J. Mol. Biol. 1983, (166) pg 557... it may also be to your advantage to lift and screen colonies before picking them.

good luck...

'Les raisonables ont dure, les passiones ont vecu'

From ldeg at midway.uchicago.edu Fri Oct 18 23:19:37 1996 From: ldeg at midway.uchicago.edu (Linda Degenstein)

Subject: Transgenic Facility Yellow Pages Message-ID: <[email protected]>

I am often contacted by people outside my university inquiring as to whether or not we can produce KO or transgenic mice for them. These are often from investigators out of the state or even out of the country. I try to match them up with an institution in their location if I know of one.

Thank you for your help. Linda Degenstein Director UCCRC Transgenic/ES Cell Facility The University of Chicago [email protected]

From pinkert at cmed.bhs.uab.edu Fri Oct 18 18:31:33 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert)

> Date: Fri, 18 Oct 1996 17:19:37 -0500 (CDT) > To: [email protected], [email protected] > From: [email protected] (Linda Degenstein) > Subject: Transgenic Facility Yellow Pages > Reply-to: [email protected]

> I am often contacted by people outside my university inquiring as to > whether or not we can produce KO or transgenic mice for them. These are > often from investigators out of the state or even out of the country. I try > to match them up with an institution in their location if I know of one. > > I would like to make a list of all the known facilities, with pertinent > info, such as institution, location, services offered, contact person, > telephone and e-mail listings I think you have info for the NICHD facility at UAB. I'm not sure if such a listing of all facilities will be problematic with the patent positions that DNX (as well as GenPharm, Lexicon, etc. for ES cell work) are staking out. But so long as the caution is there in advance, it may all be OK.

From a.annala at ucl.ac.uk Sun Oct 20 14:32:43 1996 From: a.annala at ucl.ac.uk (Dr. Alexander J. Annala)

Subject: Transgenic Facility Yellow Pages Message-ID:

Dr. Pinkert,

I am curious about the patent claims you mention. We do some pronuclear microinjection of DNA, ES cell injection into blastocysts, and aggregation of ES cells with morula to produce transgenic mice. Are individuals and/or organizations claiming patent rights which would preclude carrying out these basic transgenic procedures in academic laboratories? Are license fees to obtain the right to perform these procedures anywhere within the reach of small academic laboratories?

Are the basic transgenic patent claims analogous to the original Stanford recombinant DNA claims where a $10,000 fee was required for any lab to perform basic molecular biology techniques? It would seem few, if any, academic laboratories pay this fee today.

It might be interesting for people to have an up to date listing available on the network containing a summary of transgenic relevant patent claims and licensing arrangements/fees.

Thanks,

Alexander J. Annala, Ph.D. Wellcome Senior Research Fellow Laboratory for Molecular Pharmacology University College London Gower Street London WC1E 6BT

Tel: +44(171)380-7857 Fax: +44(171)380-7245 Email: [email protected]

>> Date: Fri, 18 Oct 1996 17:19:37 -0500 (CDT) >> To: [email protected], [email protected] >> From: [email protected] (Linda Degenstein) >> Subject: Transgenic Facility Yellow Pages >> Reply-to: [email protected] > >> I am often contacted by people outside my university inquiring as to >> whether or not we can produce KO or transgenic mice for them. These are >> often from investigators out of the state or even out of the country. I try >> to match them up with an institution in their location if I know of one. >> >> I would like to make a list of all the known facilities, with pertinent >> info, such as institution, location, services offered, contact person, >> telephone and e-mail listings >I think you have info for the NICHD facility at UAB. I'm not sure if >such a listing of all facilities will be problematic with the patent >positions that DNX >(as well as GenPharm, Lexicon, etc. for ES cell work) are staking >out. But so long as the caution is there in advance, it may all be OK. > >Carl From pinkert at cmed.bhs.uab.edu Sun Oct 20 10:02:00 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert)

> Dr. Pinkert, > > I am curious about the patent claims you mention. We do some pronuclear > microinjection of DNA, ES cell injection into blastocysts, and aggregation > of ES cells with morula to produce transgenic mice. Are individuals and/or > organizations claiming patent rights which would preclude carrying out > these basic transgenic procedures in academic laboratories? If you are providing services in house, there should be no problem. If you provide services outside your home institution, you can be liable to the folks holding patents on all the microinjection/gene transfer patents that have been awarded. Are license > fees to obtain the right to perform these procedures anywhere within the > reach of small academic laboratories? When it is allowed by the patent holders. But if the work is in-house AND for non-commercial basic research - most of the companies with the rights will leave you alone. It is only when you offer a service or begin toward commercialization that you will find yourself in need of appropriate legal counsel. > > Are the basic transgenic patent claims analogous to the original Stanford > recombinant DNA claims where a $10,000 fee was required for any lab > to perform basic molecular biology techniques? It would seem few, if > any, academic laboratories pay this fee today. Agreed, but as an example, for DNA microinjection outside of your home institution, DNX would not allow you to provide the service (as it competed with their program - but in-house basic research was automatically exempted). Now, if you want to commercialize or transfer animals, they have a basic fee structure in mind. However, if you agree to provide microinjection services to say another institution - even at/below your actual costs, you would likely be liable for treble damages . . . > > It might be interesting for people to have an up to date listing available on > the network containing a summary of transgenic relevant patent claims > and licensing arrangements/fees. I agree, but some of the ES cell patents as well as more recent targeting/expression patents are in process and disclosure is not readily available. > Good luck.

Carl A. Pinkert

From stewarv at cesmtp.ccf.org Mon Oct 21 16:27:16 1996 From: stewarv at cesmtp.ccf.org (Valerie Stewart MS)

Subject: Transgenic Facility Yellow Pages -Reply Message-ID:

Hi Linda--

Good idea, though with the patent battles, our reach is limited. Our policy so far has been to make mice for Clinic investigators and their collaborators. Here is the information you requested:

Transgenic/IKnockout Core Facility Cleveland Clinic Research Institute FF6-50, 9500 Euclid Avenue Cleveland, OH 44106 Director, Valerie Stewart Technician, Christine Brant phone (216)444-8521 fax (216)445-6257 e-mail "[email protected]" Services include pronuclear injection, blastocyst injection, cryopreservation by vitrification, training in all techniques. We also do microinjection of proteins, etc. into cell lines via an Eppendorf automated system. Services and prices can also be accessed through the Clinic's web page at "www.ccf.org". Go to Main Menu, then Research Institute, then Scientific Support Services, then Core Services.

>>> Linda Degenstein - 10/18/96 6:19 PM >>> I am often contacted by people outside my university inquiring as to whether or not we can produce KO or transgenic mice for them. These are often from investigators out of the state or even out of the country. I try to match them up with an institution in their location if I know of one.

Linda Degenstein Director UCCRC Transgenic/ES Cell Facility The University of Chicago [email protected] From YOCKEY at DCSMSERVER.MED.SC.EDU Tue Oct 22 00:15:02 1996 From: YOCKEY at DCSMSERVER.MED.SC.EDU (Courtland E. Yockey)

Subject: selection cassette-dependent phenotypes Message-ID: <[email protected]>

Hello,

Courtland Yockey ------Courtland Yockey, M.S. from the lab of Noriko Shimizu, Ph.D.

University of South Carolina School of Medicine Department of Developmental Biology & Anatomy Columbia, SC 29208 USA

Phone# 803-733-1503 Fax# 803-733-1533

From Deb_Gumucio.ANATOMY at mailgw.surg.med.umich.edu Tue Oct 22 12:30:59 1996 From: Deb_Gumucio.ANATOMY at mailgw.surg.med.umich.edu (Deb Gumucio)

Subject: selection cassette-depen Message-ID:

Reply to: RE>selection cassette-dependent phenotypes One example is in the beta globin cluster. In 1992, Mark Groudine's lab showed that insertion of a neo gene (in the Locus Control Region or LCR) upstream from the five clustered globin genes led to severe down-regulation of the downstream genes (Genes and Development 6:928). In this case, the selectable marker is not in the genes themselves, but in a regulatory region that is important for expression of the genes. In more recent work, the same lab has inserted neo in place of two of the cores of DNaseI hypersensitive sites that comprise the LCR (Genes and Development 9:2203, 1995 and Mol. Cell Biol. 16:2906, 1996). In each case, the insertion of neo in place of the core produced severe effects on globin gene expression, but removing the neo again (leaving a core deletion) restored a significant level of globin gene expression (i.e., it was worse having the neo in than having only the deletion itself). The globin cluster may be an unusual case because of the dependence of five clustered genes on a remote upstream regulatory unit (the LCR); the polar nature of the LCR-promoter interaction appears to be disrupted by the presence of the ectopic promoter on neo.

------Date: 10/21/96 6:18 PM To: Deb Gumucio From: [email protected] Hello,

Courtland Yockey ------Courtland Yockey, M.S. from the lab of Noriko Shimizu, Ph.D. University of South Carolina School of Medicine Department of Developmental Biology & Anatomy Columbia, SC 29208 USA

Phone# 803-733-1503 Fax# 803-733-1533

------RFC822 Header Follows ------Received: by mailgw.surg.med.umich.edu with SMTP;21 Oct 1996 18:16:16 -0400 Received: from majordom by bowmore.cc.ic.ac.uk with smtp (Exim 0.55 #1) id E0vFSdt-0000b0-00; Mon, 21 Oct 1996 23:16:17 +0100 Received: by ic.ac.uk (bulk_mailer for ic.ac.uk v1.6); Mon, 21 Oct 1996 23:16:12 +0100 Received: from majordom by bowmore.cc.ic.ac.uk with local (Exim 0.55 #1) id E0vFSdm-0000aR-00; Mon, 21 Oct 1996 23:16:10 +0100 Received: from juliet.ic.ac.uk [155.198.5.4] by bowmore.cc.ic.ac.uk with smtp (Exim 0.55 #1) id E0vFSdh-0000aK-00; Mon, 21 Oct 1996 23:16:05 +0100 Received: from VM.SC.EDU [129.252.41.4] by juliet.ic.ac.uk with smtp (Exim 0.55 #4) id E0vFSdf-0001kL-00; Mon, 21 Oct 1996 23:16:04 +0100 Received: from dcsmserver.med.sc.edu by VM.SC.EDU (IBM VM SMTP V2R3) with TCP; Mon, 21 Oct 96 18:15:59 EDT Received: from DCSMSERVER/SpoolDir by dcsmserver.med.sc.edu (Mercury 1.21); 21 Oct 96 18:16:16 GMT+5 Received: from SpoolDir by DCSMSERVER (Mercury 1.21); 21 Oct 96 18:15:41 GMT+5 From: "Courtland E. Yockey" Organization: USC School of Medicine To: [email protected] Date: Mon, 21 Oct 1996 18:15:02 EST Subject: selection cassette-dependent phenotypes Priority: normal X-mailer: Pegasus Mail/Mac (v2.1.2) Message-ID: <[email protected]> Sender: [email protected] Precedence: bulk Reply-To: [email protected]

From dolle at titus.u-strasbg.fr Tue Oct 22 18:19:24 1996 From: dolle at titus.u-strasbg.fr (Pascal DOLLE)

Subject: selection cassette-dependent phenotypes Message-ID: <[email protected] >

>Hello, > >I have a question regarding the phenotypes of targeted mutant mice. > >One justification for flanking selection/mutation cassettes with loxP >sites is so that the cassette - and the transcription unit associated >with the selectable marker - can be removed in order to avoid an >artifactual mutant phenotype that is dependent upon its presence. > >Does anyone know of published cases in which the phenotype of a mutant >mouse is influenced by the presence of the selectable marker itself >within the mutant locus? Cases in which the phenotype was compared, for >instance, before and after Cre-mediated excision of a loxP-flanked >cassette? > >Thanks for the information. > >Courtland Yockey >------

Dear Courtland, a couple of years ago, we have reported that some 'weird' & dominant phenotypes in chimeras harboring a Hoxd-10 disruption may have resulted from integration effect of the neo cassette in the locus. At that time, the loxP technique was not developed, so we didn't have this mutation without the selection cassette. The ref is Rijli et al., Dev. Dynamics 201, p. 366 (1994).

I guess there will be more and more cases of mutations with or without neo cassettes coming out in the next future. You could contact Filippo Rijli in our institute who may have other examples of lines with or without TK- neo integrations.

Sincerely,

Pascal DOLLE, M.D., Ph.D.

CNRS, Unite 184 INSERM I.G.B.M.C. Parc d'innovation B.P. 163 67404 ILLKIRCH Cedex FRANCE

Tel: (33) 88 65 33 34 or 40 Fax: (33) 88 65 32 01 From dbowtell at petermac.unimelb.edu.au Tue Oct 22 23:50:38 1996 From: dbowtell at petermac.unimelb.edu.au (David Bowtell)

Subject: selection cassette-dependent phenotypes In-Reply-To: <[email protected] > Message-ID:

As a follow on to the discussion about selectable markers we have placed a lacZneo fusion in frame downstream of the Sos1 gene. The marker is driven by the Sos1 promoter and there is no heterologous promoter sequences inserted. We appear to see, however, mosaic expression of lacZ in some heterozygous animals and wondered about silencing of the targeted locus in some tissues. Has anyone else encountered this?

From anna at screams.gdb.org Wed Oct 23 14:29:45 1996 From: anna at screams.gdb.org (Anna Anagnostopoulos)

Hello everyone,

Dr. Yasuzawa is trying to rid his mice of a Mycoplasma infection. I am sure he would appreciate additional suggestions.

Thanks a lot

Anna V. Anagnostopoulos

> The reason why I noticed is that my mice are infected by mycoplasma. > Especially, the clinical signs of old mice are more serious than young. > I'm worrying about both sudden death and expansion of contamination. > I already start to feed tetracyclin to them. But, the condition has not > improved yet. > If you give me any suggestion how to rescue them, I'm very happy. > > > yours sincerely, > > Kayoko Yasuzawa > > > > >

From milstone at rascal.med.harvard.edu Wed Oct 23 17:26:12 1996 From: milstone at rascal.med.harvard.edu (David Milstone)

Subject: selection cassette-dependent phenotypes Message-ID:

>Does anyone know of published cases in which the phenotype of a mutant >mouse is influenced by the presence of the selectable marker itself >within the mutant locus? Cases in which the phenotype was compared, for >instance, before and after Cre-mediated excision of a loxP-flanked >cassette? >> >Courtland Yockey

Selectable markers have been shown to influence the phenotype of mutant mice by cis-mediated effects on transciptional regulation (published examples include beta globin LCR element and CD11b). But this is presumably distinct from effects of the coding region sequences of the selectable marker itself.

David S. Milstone Vascular Research Division Department of Pathology Brigham & Women's Hospital LMRC 421 221 Longwood Avenue Boston, MA USA

From r-carver at nimr.mrc.ac.uk Thu Oct 24 17:40:21 1996 From: r-carver at nimr.mrc.ac.uk (Rick Carver)

Subject: Mycoplasma Infection Message-ID: <[email protected]>

Dear All

>Dr. Yasuzawa is trying to rid his mice of a Mycoplasma infection. I am >sure he would appreciate additional suggestions. > >> The reason why I noticed is that my mice are infected by mycoplasma. >> Especially, the clinical signs of old mice are more serious than young. >> I'm worrying about both sudden death and expansion of contamination. >> I already start to feed tetracyclin to them. But, the condition has not >> improved yet.

My advice would be to rederive them, preferably by embryo transfer. Antibiosis is a poor option because you can never be certain that it has been fully effective and you run a major risk of producing tetracycline-resistant Mycoplasma. The NRC's 'Infectious Diseases of Mice and Rats' has a nice description of the protocol to be followed post-caesarean rederivation. This protocol is equally valid if you rederive by embryo transfer. ET is much more microbiologically secure and if he is doing transgenic work he should already have the necessary equipment and skills available.

I hope that this is of assistance.

Rick Carver Head of Biological Services and Institute Veterinarian National Institute for Medical Research The Ridgeway MILL HILL London E-Mail: [email protected] NW7 1AA Tel: + 44 (0)181 959 3666 ext 2199 United Kingdom Fax: + 44 (0)181 913 8601 From dknudsen at scruznet.com Thu Oct 24 06:16:18 1996 From: dknudsen at scruznet.com (Dave Knudsen)

>Hello everyone, > >Dr. Yasuzawa is trying to rid his mice of a Mycoplasma infection. I am >sure he would appreciate additional suggestions. > >Thanks a lot > >Anna V. Anagnostopoulos > >> The reason why I noticed is that my mice are infected by mycoplasma. >> Especially, the clinical signs of old mice are more serious than young. >> I'm worrying about both sudden death and expansion of contamination. >> I already start to feed tetracyclin to them. But, the condition has not >> improved yet. >> If you give me any suggestion how to rescue them, I'm very happy.

Anna -

I would agree with Rick Carver in that rederivation is the best way to proceed for a contaminated colony. The assumption under this agreement is that the affected colony is a small breeding colony, and not a group of stock animals that could be eradicated and replaced at the end of an experiment. I also assume that your colleague, Dr. Yasuzawa, has completed the diagnostic process and confirmed mycoplasmosis as the problem. This diagnosis can be made by serology, culture, or histopathology (preferably all three for a colony, to reduce the chance of false negative findings). The confirmation itself is important to resolution of the problem - I can think of several instances where other opportunistic pathogens, environmental factors, immunodeficiencies, and/or transgenic phenotypes have mimicked the clinical presentation of murine mycoplasmosis quite closely. Also, I would add that antibiosis may help the clinical appearance of the animals by temporarily arresting disease progression, but it will do nothing in evoking a cure, and treated animals will continue to be carriers to infect new introductions (and births) into the colony.

I would personally be most comfortable, however, in dealing with the rederivation issue by caesarian section; I suspect that confidence is gained from the method most familiar. Over the years I have had more positive experiences with caesarian rederivation in eliminating pathogens from mouse colonies, and embryo or ovarian manipulations seem always to break at some point. Perhaps the best method could be determined by close questioning of the veterinary staff of the affected institution and working with them to resolve the outbreak.

Dave Knudsen DVM, DACLAM Scotts Valley CA, USA

1996 November

On Mon, 25 Nov 1996, Nick Carboni wrote:

> Hi, > > A student mailed me these questions ... maybe one of you could help him by > answering these questions or advising him of some references. > > >1- About cytoplasmic microinjection using polylysine/DNA mixture: How > >polylysine provides cytoplasmic microinjection? Has anyone ever tried it? > Is >this technique really revolutionary? > > > >2- Do some of you consider it possible to product transgenic mice by random > >insertion using ES cells ? > > > >3- What about retroviral infection efficiency in the production of > transgenic & >Ko mice > > > Thank you in advance > > Nick > ------> Nick Carboni, VP > Valoritech > 1253, McGill College Av., room 620 > Montreal, Quebec, Canada > H3B 2Y5 > Tel.: (514) 866-8271 / Fax: (514) 866-8272 > E-Mail: [email protected] > > > > From pinkert at cmed.bhs.uab.edu Tue Nov 26 08:29:43 1996 From: pinkert at cmed.bhs.uab.edu (Dr. C.A. Pinkert)

Subject: Technical questions Message-ID: <19961126143625396.AAA130@cap1>

> A student mailed me these questions ... maybe one of you could help him by > answering these questions or advising him of some references. > > >1- About cytoplasmic microinjection using polylysine/DNA mixture: How > >polylysine provides cytoplasmic microinjection? Has anyone ever tried it? Is >this technique really revolutionary? It was published - see RL Page et al., Transgenic Research 4:353 (1995). > >2- Do some of you consider it possible to product transgenic mice by random >insertion using ES cells ? It is doable - but at issue is whether it is cost-effective compared to direct DNA microinjection (dependent upon the particular lab ...)?

>3- What about retroviral infection efficiency in the production of > transgenic & Ko mice It isn't advantageous - and gene expression issues are not inconsequential. You can find a goodly number of reviews on the subject (relative efficiencies, etc.) with a Medline or comparable abstracting system.

Carl A. Pinkert

From McDonald at WSUHUB.UC.TWSU.EDU Wed Nov 27 14:05:46 1996 From: McDonald at WSUHUB.UC.TWSU.EDU (J. D. McDonald)

Subject: Noise problems and solutions Message-ID: <[email protected]>

I am the director of our university's animal care facility and I would like to call upon the mouse husbandry expertise of this group with regard to a specific noise issue. Our building will soon be undergoing an extensive upgrading of the HVAC (heating, ventilation, and air conditioning) system. This will not occur within the cluster of rooms that comprise our animal care facility but will be occurring all around it for a period of about a month. We are anticipating significant amounts of intermittent construction noise and are investigating ways of dealing with this so as to minimize the stress levels on our research animals (primarily mice). In discussing options, one idea is to introduce some type of white noise to mask the construction noise. Does anyone out there have any experience with this strategy? Can you point me to pertinent literature (a quick Medline search didn't turn up anything of apparent value)? Do you know of other more effective ways of dealing with this problem? Please send any advice that will likely be helpful by email. If I get a good number of responses, I will post a summary.

J. David McDonald, Ph. D., Assistant Professor Wichita State University Department of Biological Sciences 1845 Fairmount, Box #26 Wichita, KS 67260-0026 Telephone 316-978-3111 x16 Fax 316-978-3772 Voicemail 316-978-3978 x6097 Email [email protected]

From crussell at ResGen.COM Wed Nov 27 16:11:47 1996 From: crussell at ResGen.COM (Chris Russell)

Subject: Technical questions Message-ID: <[email protected]>

Further to Dr. Pinkert's answers to the following question:

>>1- About cytoplasmic microinjection using polylysine/DNA mixture: How >>polylysine provides cytoplasmic microinjection? Has anyone ever tried it? >Is >this technique really revolutionary?

R. L. Page, S. P. Butler, A. Subramanian, F. C. Gwazdauskas, J. L. Johnson, and W. H. Velander. 1995. "Transgenesis in Mice by Cytoplasmic Injection of Polylysine/DNA Mixtures". Transgenic Research 4:353-360

The polyLysine-DNA (pLDNA) mixture is injected into the cytoplasm of fertilized embryos, similar to the technique of pronuclear microinjection. However, in this case, it is not necessary to hit nor even see the pronucleus. The technique is not a efficient as pronuclear microinjection -- about 15% maximum transgenesis observed in mice -- but does give rise to functional, expressing transgenic animals, which seemed to be equivalent to transgenic mice made by pronuclear microinjection. It also provides another technique in the arsenal of transgenic animal production. Is the technique revolutionary? Depends on your opinion of demonstration of new techniques. Apparently the US patent office thought so. The patent was awarded to Page, Velander, and perhaps one other author. To the best of my knowledge, PPL, Inc (of Blacksburg, VA and Edinburgh, Scotland) holds the exclusive license to cytoplasmic injection for generation of transgenic animals.

Let me know if there are further questions, and I'd be glad to try to help (I was in the transgenic research group at Virginia Tech at the time of development of the technique).

Chris *********************************************************************** **** Christopher G. Russell, Ph.D. RESEARCH GENETICS, INC Resources for Research -_+ _+_+ ++-_ _+-_ _+_ _++ _+-_ -+ -+ -+ +- +- -+ + -+ -+ -+ + -+ -+ -+ -+ +- -+ -+ -+ -+ -+ -+ -+ -+ -++-+ -+_-+ -+_+ -++- -+_++ -+_+ 2700 Memorial Parkway SW Huntsville, AL 35801 Phone 1-800-533-4363, ext 2211 or Direct 1-800-711-2089 U.K. 0-800-89-1393 Fax 1-205-551-1021 or 1-205-536-9016 [email protected] *********************************************************************** ******

From tonyjames at cuhk.edu.hk Fri Nov 29 02:40:51 1996 From: tonyjames at cuhk.edu.hk (A E James)

Subject: Noise problems and solutions Message-ID: <[email protected]>

Can I suggest you correspond with Dr. Gerald Glough in the UK, he is a recognised expert on these matters. A relevant paper is Enviromental Effects on Animals used in Biomedical Research. G. Clough, Biol. Rev. Vol. 57,pp. 487-523 Gerald can be contacted through Alanan Consultancy Sevives PO Box 230 York YO1 1GG England, ph/fx 44-1904-656368 >I am the director of our university's animal care facility and I would like >to call upon the mouse husbandry expertise of this group with regard to a >specific noise issue. Our building will soon be undergoing an extensive >upgrading of the HVAC (heating, ventilation, and air conditioning) system. >This will not occur within the cluster of rooms that comprise our animal >care facility but will be occurring all around it for a period of about a >month. We are anticipating significant amounts of intermittent construction >noise and are investigating ways of dealing with this so as to minimize the >stress levels on our research animals (primarily mice). In discussing >options, one idea is to introduce some type of white noise to mask the >construction noise. Does anyone out there have any experience with this >strategy? Can you point me to pertinent literature (a quick Medline search >didn't turn up anything of apparent value)? Do you know of other more >effective ways of dealing with this problem? Please send any advice that >will likely be helpful by email. If I get a good number of responses, I >will post a summary. > >J. David McDonald, Ph. D., Assistant Professor >Wichita State University >Department of Biological Sciences >1845 Fairmount, Box #26 >Wichita, KS 67260-0026 >Telephone 316-978-3111 x16 >Fax 316-978-3772 >Voicemail 316-978-3978 x6097 >Email [email protected] > > > > Tony James BVSc MACVSc Director Laboratory Animal Unit Chinese University of Hong Kong Shatin, New Territories, Hong Kong ph: 852 2609 6862 fx: 852 2603 5723 email: [email protected] ...... "nothing great was achieved without enthusiasm." R. W. Emerson 1996 December

From pasceri at sickkids.on.ca Mon Dec 2 17:05:25 1996 From: pasceri at sickkids.on.ca (Peter Pasceri) Date: Wed Aug 20 11:52:56 2003 Subject: Seasonal changes Message-ID:

I have a question which is not pressing but would appreciate any input from those of you that have experienced the same.

Over the past three years I have noticed that my animals seem to behave differently during the months of November and December. They give less eggs when superovulated and there seems to be more resorbtions than is acceptable. Last week more than 80% resorbtions !

Culture media supports growth in vitro and is not suspected. Things usually straighten out by late January.

Any ideas?

-- Peter Pasceri [email protected] Genetics Research The Hospital for Sick Children Toronto, Ontario,Canada Phone: ( 416 ) 813-8169

From gotto at leland.Stanford.EDU Mon Dec 2 16:44:22 1996 From: gotto at leland.Stanford.EDU (Glen Otto) Date: Wed Aug 20 11:52:56 2003 Subject: Seasonal changes In-Reply-To: Message-ID:

>Over the past three years I have noticed that my animals seem to behave >differently during the months of November and December. They give less >eggs when superovulated and there seems to be more resorbtions than is >acceptable. Last week more than 80% resorbtions ! Statistically significant annual cycles in mouse reproductive efficiency (even in climate-controlled indoor facilities) is not uncommon in populations large enough to notice it, and I've been told that some major breeding facilities take it into account when making production projections.

I'm not sure anybody knows why, and when I mentioned it to a prominent sociobiologist a few years ago she thought it would make a great thesis study. Maybe a fallback project for someone who can't get a strain made because the mice don't produce! glen

______Glen Otto, DVM Dept. of Comparative Medicine [email protected] Stanford University Quad 7, Bldg. 330 415/723-3876 voice Stanford, CA 94305-5410 415/725-0940 fax

From jklohse at facstaff.wisc.edu Mon Dec 2 22:08:33 1996 From: jklohse at facstaff.wisc.edu (Jan K. Lohse) Date: Wed Aug 20 11:52:56 2003 Subject: Seasonal changes In-Reply-To: Message-ID:

Many workers with rodent embryos experience some seasonal variation. This is sometimes a slump in early winter, as you described, and sometimes one problem period around October & another around April, when the weather is changing the most. This occurs even in light/temperature/humidity controlled colonies. I have never heard of a good, conclusive study tracking down the reasons for this, but it seems to be fairly common.

From browng at medicine.wustl.edu Tue Dec 3 07:09:24 1996 From: browng at medicine.wustl.edu (Gary Brown) Date: Wed Aug 20 11:52:56 2003 Subject: Seasonal changes In-Reply-To: Message-ID:

Hi! Just my $0.02 worth... I too have experienced such seasonal phenomena in the past, and although might give a probable cause I can't pretend the answer will be easy to implement! In general, the most difficult times of year coincide with the late phase of exterior climactic change ie Fall and Spring. The building in which animals are housed will undergo its own climactic adjustments correspondingly as air conditioning, humidity control and heating readjust. The efficiency (often times the age) of the building in making this transition will determine the severity of the observed effect as a new environmental stress is added to post surgical mice at this time. All I can suggest is that you work with your building manager / facility manager to minimize the impact of such changes, and maintain accurate records of high/low humidity, temp etc. to visualise if this is indeed the problem.

Another small(?) consideration. In my own situation, we were originally housed in the basement area of the local Childrens Hospital until very recently - we've since moved to a purpose built facility :-) In this situation, the environmental controls were shared between our mouse floor and the one above (a "people" floor), necessitating a less-than- desirable compromise. Nothing's ever easy......

Regards,

On Mon, 2 Dec 1996, Peter Pasceri wrote:

> > I have a question which is not pressing but would appreciate any input from > those of you that have experienced the same. > > Over the past three years I have noticed that my animals seem to behave > differently during the months of November and December. They give less > eggs when superovulated and there seems to be more resorbtions than is > acceptable. Last week more than 80% resorbtions ! > > Culture media supports growth in vitro and is not suspected. Things > usually straighten out by late January. > > Any ideas? > > Peter > > > -- > Peter Pasceri > [email protected] > Genetics Research > The Hospital for Sick Children > Toronto, Ontario,Canada > Phone: ( 416 ) 813-8169 > > > > >

From tjf at uci.edu Tue Dec 3 11:43:34 1996 From: tjf at uci.edu (Tom Fielder) Date: Wed Aug 20 11:52:56 2003 Subject: in vitro vs. in vivo expression Message-ID:

I would like to get some feedback from those of you involved in transgenic research (this will be posted on Embryo Mail and Compmed as well). I am running a new transgenic core facility at UC-Irvine. We are debating whether or not to require some evidence of protein expression from DNA constructs before doing pronuclear injection. On the one hand, it would be nice to know that the construct has at least some chance of working and that the investigator didn't make a mistake in the engineering. On the other hand, some people argue that in vitro expression cannot guarantee in vivo expression (certainly true) and is in fact not even a decent predictor of in vivo success. What do people require in terms of construct testing (e.g., RE digest, gel picture, sequencing, in vitro expression), and does anyone have any statistics on the predictability of in vivo expression from in vitro results? If I get enough responses, I'll summarize for the list. Thanks for your help! Tom

Thomas J. Fielder UC-Irvine University Lab Animal Resources 147 BSA Irvine, CA 92697-1310 e-mail: [email protected] phone: 714-824-8579 FAX: 714-824-2003 From david.threadgill at mcmail.vanderbilt.edu Thu Dec 5 10:00:49 1996 From: david.threadgill at mcmail.vanderbilt.edu ([email protected]) Date: Wed Aug 20 11:52:56 2003 Subject: spretus Message-ID: <[email protected]>

Does anyone have experience with superovulating spretus females? Do they respond to the same level of hormones as musculus? Do spretus males mate reliably with superovulated females? Thanks, David Threadgill Dept of Cell Biology Vanderbilt Univ Nashville, TN [email protected]

From p.sobieszczuk at ic.ac.uk Fri Dec 6 17:40:36 1996 From: p.sobieszczuk at ic.ac.uk (Peter Sobieszczuk) Date: Wed Aug 20 11:52:56 2003 Subject: Seasonal changes/Forwarded from Pete Willan Message-ID:

>From: Pete Willan >To: [email protected] >Date: Thu, 5 Dec 1996 09:49:20 +0100 (BST) >Subject: Re: Seasonal changes >Reply-to: "Division of Biomedical Services"@leicester.ac.uk, > [email protected] >CC: [email protected] >Priority: normal >X-mailer: Pegasus Mail for Windows (v2.40) >Message-ID: <[email protected]> > >Hi to all > >This may not be the most authentic answer to reduction in breeding >colonies and embryo reabsorbtion(I am assuming that changes in time clocks >have also been investigated) has anyone out there >looked at the following senario... > > >The only variation into the breeding colony that may effect the >animals could be born(exuse pun) from the effect of seasonal >changes on the Animal Welfare Staff being transmitted somehow >to the animals. > >I would be interested in comments. > >Seasonal Greetings to all! > >Pete Willan

From p.sobieszczuk at ic.ac.uk Sat Dec 7 16:22:37 1996 From: p.sobieszczuk at ic.ac.uk (Peter Sobieszczuk) Date: Wed Aug 20 11:52:56 2003 Subject: Noise problems/Forwarded from [email protected] Message-ID:

>To: [email protected] >From: [email protected] (Rachael Parkinson) >Subject: Re: Noise problems >Content-Length: 718 > >My department is currently undergoing construction work and, yes, this has >had quite an adverse effect on mouse breeding and cannibalism of mouse >pups. I have no good news. We had our "temporary" mouse room >sound-proofed to try to minimalise the noise levels in there, but the >construction work has caused quite serious vibrations throughout the >building which I believe to be the primary cause of my animals' stress. If >your building works are expected to cause vibrations, then I think this >will be more of a concern than noise which may be able to be masked (by >white noise as you have suggested). > >Rachael Parkinson >Department Of Anatomy and Cell Biology >University of Melbourne >Parkville VIC 3052 >Australia.

From BIOGEN at aol.com Sat Dec 7 13:51:07 1996 From: BIOGEN at aol.com ([email protected]) Date: Wed Aug 20 11:52:56 2003 Subject: Noise problems/Forwarded from [email protected] Message-ID: <[email protected]>

My first interaction with the group is somewhat off-the-wall: In regard to Rachel's observation of the decline in breeding and the associated cannabalism as a result of the auditory assault generated from construction work-- Would you please comment on the estimated percentile of decline in breeding and/or the possible general effect this may have on the population in general? And, do you have any way of quantifying the decibel level and mean frequency of the noise?

If these seem like strange requests, upon further consideration you might see where I'm going: If the decline in breeding is statistically large, and the frequencies involved can be isolated into a range minimally obtrusive to humans, it might be of use in eradication or in the prevention of infestation in structures. As my primary education was in electronic engineering, I am always curious about the possibilities of interaction between disciplines (biology and electronics). Thanks for your consideration!

Kind Regards,

Lou Jones / Biogenics

From tjf at uci.edu Wed Dec 11 15:25:19 1996 From: tjf at uci.edu (Tom Fielder) Date: Wed Aug 20 11:52:57 2003 Subject: in vitro expression testing Message-ID:

Here is a summary of the replies I received in response to my question regarding the advisibility of in vitro expression testing of DNA constructs prior to pronuclear injection:

There were 10 respondents. Apparently no one demands mandatory in vitro testing, although 3 respondents said they ask for or encourage such testing when possible (e.g., if an appropriate cell line is available). Of the seven replies which listed what they do require, the following items were mentioned: gel photo: 6 restriction digest: 2 sequence data around cloning sites: 2 evidence of specific PCR assay: 1

Thanks very much for your replies.

Thomas J. Fielder UC-Irvine University Lab Animal Resources 147 BSA Irvine, CA 92697-1310 e-mail: [email protected] phone: 714-824-8579 FAX: 714-824-2003

From i.rosewell at icrf.icnet.uk Tue Dec 17 13:01:45 1996 From: i.rosewell at icrf.icnet.uk (Ian -Transgenic Unit) Date: Wed Aug 20 11:52:57 2003 Subject: transmission? Message-ID:

We have recently achieved germline transmission from 2 independent clones. The clones were from seperate knockout projects and seperate ES cell lines. The clones transmited poorly. In the first case after a number of litters one chimera gave rise to what was initially identified to me as a chimera, the coat colour which should have been( Bl6 x 129) agouti was uneven with areas of black fur. The chimera that gave rise to this then went on to father a true agouti and het. mouse. The first mouse was negative. This has happened again. Can anyone provide an explanation?

Thanks

From anna at screams.gdb.org Thu Dec 19 09:25:39 1996 From: anna at screams.gdb.org (Anna Anagnostopoulos) Date: Wed Aug 20 11:52:57 2003 Subject: purchase/breeding of APO E KO mice In-Reply-To: Message-ID:

On Tue, 19 Nov 1996, Frederik =?iso-8859-1?Q?Dagn=E6s=2DHansen?= wrote:

Dear Frederik,

I don't know much about availability in Europe. However, Apoe strains C57BL/6J-Apoe(tm1Unc) generated by Dr. Nobuyo Maeda and strain B6, 129-Apoe(tm1Unc) Ldlr(tm1Her) generated by Dr. Joachim Herz are both available at the Induced Mutant Resources at the Jackson Laboratories, Bar Harbor, Maine, USA (Stock #s are JR2052 and JR2245, respectively). The latter is of limited distribution. You may check the IMR Web site for additional information (http://lena.jax.org/resources/documents/imr/). I can also provide the following contact information to you through TBASE (http://www.gdb.org/Dan/tbase/tbase.html):

Dr. Nobuyo Maeda

Univ. of North Carolina at Chaperl HIll Dept of Pathology CB7525 Brinkhous-Bullitt Building Chaperl Hill, NC 29599-7525 (608) 262-1047

You may also want to contact:

Dr. Jan Breslow The Rockefeller University Biochemical Genetics and Metabolism Box 29 HOS 1230 York Ave, New York, NY 10021 (212) 570-7700

They may answer some of your questions regarding availability and breeding.

I hope this will be of some assistance

Sincerely

Anna V. Anagnostopoulos e-mail: [email protected]

> Hej everybody > > A researcher working in cardiovascular research whishes to work with the > APO E knockout mice (C57BL/6J-APO E KO ) . > As it is difficult to purchase these mice from commercial breeders in > Europe, he has tryed to establish a small breeding colony of these. > The outcome is very bad as the mothers eat the pups just after delivery. > Does anyone have some good ideas how to get these mice, how to make them > breed???????+ > > best regards > > Frederik Dagn�s-Hansen > > Frederik Dagnaes-Hansen,D.V.M., Ph.D. > Inst. Med.Microbiol.& Imm. > Bartholin Building, Building 240, > University of Aarhus, > 8000 Aarhus C, > Tel + 45 89 42 1774 > Fax + 45 8819 6128 > Email: [email protected] > > > > > >

From tjf at uci.edu Mon Dec 30 10:49:02 1996 From: tjf at uci.edu (Tom Fielder) Date: Wed Aug 20 11:52:57 2003 Subject: Balb/C knockouts Message-ID:

Does anyone know of a source of CD4 and/or CD8 knockouts in Balb/C mice? A colleague of mine has contacted Taconic and Jax and they only have them in C57's. Thanks in advance. Tom

From pasceri at sickkids.on.ca Mon Dec 2 21:05:25 1996 From: pasceri at sickkids.on.ca (Peter Pasceri)

Subject: Seasonal changes Message-ID:

I have a question which is not pressing but would appreciate any input from those of you that have experienced the same.

Over the past three years I have noticed that my animals seem to behave differently during the months of November and December. They give less eggs when superovulated and there seems to be more resorbtions than is acceptable. Last week more than 80% resorbtions !

Culture media supports growth in vitro and is not suspected. Things usually straighten out by late January.

Any ideas?

-- Peter Pasceri [email protected] Genetics Research The Hospital for Sick Children Toronto, Ontario,Canada Phone: ( 416 ) 813-8169

From gotto at leland.Stanford.EDU Tue Dec 3 00:44:22 1996 From: gotto at leland.Stanford.EDU (Glen Otto)

Subject: Seasonal changes In-Reply-To: Message-ID:

>Over the past three years I have noticed that my animals seem to behave >differently during the months of November and December. They give less >eggs when superovulated and there seems to be more resorbtions than is >acceptable. Last week more than 80% resorbtions !

Statistically significant annual cycles in mouse reproductive efficiency (even in climate-controlled indoor facilities) is not uncommon in populations large enough to notice it, and I've been told that some major breeding facilities take it into account when making production projections.

I'm not sure anybody knows why, and when I mentioned it to a prominent sociobiologist a few years ago she thought it would make a great thesis study. Maybe a fallback project for someone who can't get a strain made because the mice don't produce! glen

______Glen Otto, DVM Dept. of Comparative Medicine [email protected] Stanford University Quad 7, Bldg. 330 415/723-3876 voice Stanford, CA 94305-5410 415/725-0940 fax From jklohse at facstaff.wisc.edu Tue Dec 3 05:08:33 1996 From: jklohse at facstaff.wisc.edu (Jan K. Lohse)

Many workers with rodent embryos experience some seasonal variation. This is sometimes a slump in early winter, as you described, and sometimes one problem period around October & another around April, when the weather is changing the most. This occurs even in light/temperature/humidity controlled colonies. I have never heard of a good, conclusive study tracking down the reasons for this, but it seems to be fairly common.

From browng at medicine.wustl.edu Tue Dec 3 13:09:24 1996 From: browng at medicine.wustl.edu (Gary Brown)

Just my $0.02 worth... I too have experienced such seasonal phenomena in the past, and although might give a probable cause I can't pretend the answer will be easy to implement! In general, the most difficult times of year coincide with the late phase of exterior climactic change ie Fall and Spring. The building in which animals are housed will undergo its own climactic adjustments correspondingly as air conditioning, humidity control and heating readjust. The efficiency (often times the age) of the building in making this transition will determine the severity of the observed effect as a new environmental stress is added to post surgical mice at this time. All I can suggest is that you work with your building manager / facility manager to minimize the impact of such changes, and maintain accurate records of high/low humidity, temp etc. to visualise if this is indeed the problem.

Another small(?) consideration. In my own situation, we were originally housed in the basement area of the local Childrens Hospital until very recently - we've since moved to a purpose built facility :-) In this situation, the environmental controls were shared between our mouse floor and the one above (a "people" floor), necessitating a less-than- desirable compromise. Nothing's ever easy......

Regards,

On Mon, 2 Dec 1996, Peter Pasceri wrote:

> > I have a question which is not pressing but would appreciate any input from > those of you that have experienced the same. > > Over the past three years I have noticed that my animals seem to behave > differently during the months of November and December. They give less > eggs when superovulated and there seems to be more resorbtions than is > acceptable. Last week more than 80% resorbtions ! > > Culture media supports growth in vitro and is not suspected. Things > usually straighten out by late January. > > Any ideas? > > Peter > > > -- > Peter Pasceri > [email protected] > Genetics Research > The Hospital for Sick Children > Toronto, Ontario,Canada > Phone: ( 416 ) 813-8169 > > > > >

From tjf at uci.edu Tue Dec 3 19:43:34 1996 From: tjf at uci.edu (Tom Fielder)

Subject: in vitro vs. in vivo expression Message-ID:

I would like to get some feedback from those of you involved in transgenic research (this will be posted on Embryo Mail and Compmed as well). I am running a new transgenic core facility at UC-Irvine. We are debating whether or not to require some evidence of protein expression from DNA constructs before doing pronuclear injection. On the one hand, it would be nice to know that the construct has at least some chance of working and that the investigator didn't make a mistake in the engineering. On the other hand, some people argue that in vitro expression cannot guarantee in vivo expression (certainly true) and is in fact not even a decent predictor of in vivo success. What do people require in terms of construct testing (e.g., RE digest, gel picture, sequencing, in vitro expression), and does anyone have any statistics on the predictability of in vivo expression from in vitro results? If I get enough responses, I'll summarize for the list. Thanks for your help! Tom

From david.threadgill at mcmail.vanderbilt.edu Thu Dec 5 16:00:49 1996 From: david.threadgill at mcmail.vanderbilt.edu ([email protected])

Subject: spretus Message-ID: <[email protected]>

Does anyone have experience with superovulating spretus females? Do they respond to the same level of hormones as musculus? Do spretus males mate reliably with superovulated females? Thanks, David Threadgill Dept of Cell Biology Vanderbilt Univ Nashville, TN [email protected] From p.sobieszczuk at ic.ac.uk Fri Dec 6 17:40:36 1996 From: p.sobieszczuk at ic.ac.uk (Peter Sobieszczuk)

Subject: Seasonal changes/Forwarded from Pete Willan Message-ID:

>From: Pete Willan >To: [email protected] >Date: Thu, 5 Dec 1996 09:49:20 +0100 (BST) >Subject: Re: Seasonal changes >Reply-to: "Division of Biomedical Services"@leicester.ac.uk, > [email protected] >CC: [email protected] >Priority: normal >X-mailer: Pegasus Mail for Windows (v2.40) >Message-ID: <[email protected]> > >Hi to all > >This may not be the most authentic answer to reduction in breeding >colonies and embryo reabsorbtion(I am assuming that changes in time clocks >have also been investigated) has anyone out there >looked at the following senario... > > >The only variation into the breeding colony that may effect the >animals could be born(exuse pun) from the effect of seasonal >changes on the Animal Welfare Staff being transmitted somehow >to the animals. > >I would be interested in comments. > >Seasonal Greetings to all! > >Pete Willan

From p.sobieszczuk at ic.ac.uk Sat Dec 7 16:22:37 1996 From: p.sobieszczuk at ic.ac.uk (Peter Sobieszczuk)

Subject: Noise problems/Forwarded from [email protected] Message-ID:

>To: [email protected] >From: [email protected] (Rachael Parkinson) >Subject: Re: Noise problems >Content-Length: 718 > >My department is currently undergoing construction work and, yes, this has >had quite an adverse effect on mouse breeding and cannibalism of mouse >pups. I have no good news. We had our "temporary" mouse room >sound-proofed to try to minimalise the noise levels in there, but the >construction work has caused quite serious vibrations throughout the >building which I believe to be the primary cause of my animals' stress. If >your building works are expected to cause vibrations, then I think this >will be more of a concern than noise which may be able to be masked (by >white noise as you have suggested). > >Rachael Parkinson >Department Of Anatomy and Cell Biology >University of Melbourne >Parkville VIC 3052 >Australia.

From BIOGEN at aol.com Sat Dec 7 18:51:07 1996 From: BIOGEN at aol.com ([email protected]) Date: Fri Jul 9 14:03:00 2004 Subject: Noise problems/Forwarded from [email protected] Message-ID: <[email protected]>

My first interaction with the group is somewhat off-the-wall: In regard to Rachel's observation of the decline in breeding and the associated cannabalism as a result of the auditory assault generated from construction work-- Would you please comment on the estimated percentile of decline in breeding and/or the possible general effect this may have on the population in general? And, do you have any way of quantifying the decibel level and mean frequency of the noise?

If these seem like strange requests, upon further consideration you might see where I'm going: If the decline in breeding is statistically large, and the frequencies involved can be isolated into a range minimally obtrusive to humans, it might be of use in eradication or in the prevention of infestation in structures. As my primary education was in electronic engineering, I am always curious about the possibilities of interaction between disciplines (biology and electronics). Thanks for your consideration!

Kind Regards, Lou Jones / Biogenics

From tjf at uci.edu Wed Dec 11 23:25:19 1996 From: tjf at uci.edu (Tom Fielder) Date: Fri Jul 9 14:03:00 2004 Subject: in vitro expression testing Message-ID:

Here is a summary of the replies I received in response to my question regarding the advisibility of in vitro expression testing of DNA constructs prior to pronuclear injection:

There were 10 respondents. Apparently no one demands mandatory in vitro testing, although 3 respondents said they ask for or encourage such testing when possible (e.g., if an appropriate cell line is available). Of the seven replies which listed what they do require, the following items were mentioned: gel photo: 6 restriction digest: 2 sequence data around cloning sites: 2 evidence of specific PCR assay: 1

Thanks very much for your replies.

From i.rosewell at icrf.icnet.uk Tue Dec 17 12:01:45 1996 From: i.rosewell at icrf.icnet.uk (Ian -Transgenic Unit) Date: Fri Jul 9 14:03:00 2004 Subject: transmission? Message-ID:

We have recently achieved germline transmission from 2 independent clones. The clones were from seperate knockout projects and seperate ES cell lines. The clones transmited poorly. In the first case after a number of litters one chimera gave rise to what was initially identified to me as a chimera, the coat colour which should have been( Bl6 x 129) agouti was uneven with areas of black fur. The chimera that gave rise to this then went on to father a true agouti and het. mouse. The first mouse was negative. This has happened again. Can anyone provide an explanation?

Thanks

From anna at screams.gdb.org Thu Dec 19 14:25:39 1996 From: anna at screams.gdb.org (Anna Anagnostopoulos) Date: Fri Jul 9 14:03:00 2004 Subject: purchase/breeding of APO E KO mice In-Reply-To: Message-ID:

On Tue, 19 Nov 1996, Frederik =?iso-8859-1?Q?Dagn=E6s=2DHansen?= wrote:

Dear Frederik,

I don't know much about availability in Europe. However, Apoe strains C57BL/6J-Apoe(tm1Unc) generated by Dr. Nobuyo Maeda and strain B6, 129-Apoe(tm1Unc) Ldlr(tm1Her) generated by Dr. Joachim Herz are both available at the Induced Mutant Resources at the Jackson Laboratories, Bar Harbor, Maine, USA (Stock #s are JR2052 and JR2245, respectively). The latter is of limited distribution. You may check the IMR Web site for additional information (http://lena.jax.org/resources/documents/imr/). I can also provide the following contact information to you through TBASE (http://www.gdb.org/Dan/tbase/tbase.html):

Dr. Nobuyo Maeda

Univ. of North Carolina at Chaperl HIll Dept of Pathology CB7525 Brinkhous-Bullitt Building Chaperl Hill, NC 29599-7525 (608) 262-1047

You may also want to contact:

Dr. Jan Breslow The Rockefeller University Biochemical Genetics and Metabolism Box 29 HOS 1230 York Ave, New York, NY 10021 (212) 570-7700 They may answer some of your questions regarding availability and breeding.

I hope this will be of some assistance

Sincerely

Anna V. Anagnostopoulos e-mail: [email protected]

> Hej everybody > > A researcher working in cardiovascular research whishes to work with the > APO E knockout mice (C57BL/6J-APO E KO ) . > As it is difficult to purchase these mice from commercial breeders in > Europe, he has tryed to establish a small breeding colony of these. > The outcome is very bad as the mothers eat the pups just after delivery. > Does anyone have some good ideas how to get these mice, how to make them > breed???????+ > > best regards > > Frederik Dagn�s-Hansen > > Frederik Dagnaes-Hansen,D.V.M., Ph.D. > Inst. Med.Microbiol.& Imm. > Bartholin Building, Building 240, > University of Aarhus, > 8000 Aarhus C, > Tel + 45 89 42 1774 > Fax + 45 8819 6128 > Email: [email protected] > > > > > >

From tjf at uci.edu Mon Dec 30 18:49:02 1996 From: tjf at uci.edu (Tom Fielder) Date: Fri Jul 9 14:03:00 2004 Subject: Balb/C knockouts Message-ID:

Does anyone know of a source of CD4 and/or CD8 knockouts in Balb/C mice? A colleague of mine has contacted Taconic and Jax and they only have them in C57's. Thanks in advance. Tom