DOCSLIB.ORG

Yeast Metabolic and Signaling Genes Are Required for Heat-Shock Survival

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Yeast Metabolic and Signaling Genes Are Required for Heat-Shock Survival Yeast metabolic and signaling genes are required for PNAS PLUS heat-shock survival and have little overlap with the heat-induced genes Patrick A. Gibneya,b,1, Charles Lub,1, Amy A. Caudya,2,3, David C. Hessc,3, and David Botsteina,b,4 aLewis-Sigler Institute for Integrative Genomics and bDepartment of Molecular Biology, Princeton University, Princeton, NJ 08544; and cDepartment of Biology, Santa Clara University, Santa Clara, CA 95053 Contributed by David Botstein, September 30, 2013 (sent for review August 25, 2013) Genome-wide gene-expression studies have shown that hundreds or proteins that increase in abundance as part of the heat-shock of yeast genes are induced or repressed transiently by changes in response (1, 11–14). Classically, this set of genes/proteins includes temperature; many are annotated to stress response on this basis. the molecular chaperones, which often are induced in response to To obtain a genome-scale assessment of which genes are function- stress (14). However, by using the deletion collection to assay ally important for innate and/or acquired thermotolerance, we thermotolerance, we also can identify genes important for stress combined the use of a barcoded pool of ∼4,800 nonessential, pro- resistance that are either constitutively present or posttransla- totrophic Saccharomyces cerevisiae deletion strains with Illumina- tionally modified. We therefore carried out two kinds of exper- based deep-sequencing technology. As reported in other recent iment to examine heat-stress resistance. First we studied the 50 °C studies that have used deletion mutants to study stress responses, survival time-course of samples from a steady-state chemostat cul- we observed that gene deletions resulting in the highest thermo- ture of the entire prototrophic barcoded deletion collection. Sec- sensitivity generally are not the same as those transcriptionally ond, we tested for the acquisition of thermotolerance by shifting fi induced in response to heat stress. Functional analysis of identi ed the temperature of the chemostat to 37 °C, a temperature known genes revealed that metabolism, cellular signaling, and chromatin to suffice for induction of the heat-stress–associated gene set, regulation play roles in regulating thermotolerance and in acquired before doing another 50 °C survival time-course. Results of these thermotolerance. However, for most of the genes identified, the experiments are summarized in Table 1. molecular mechanism behind this action remains unclear. In fact, a In the first experiment, we identified a surprisingly small num- large fraction of identified genes are annotated as having unknown ber of deletion strains (55 gene deletions) that differ significantly functions, further underscoring our incomplete understanding of the response to heat shock. We suggest that survival after heat from the rest of the population in their heat sensitivity (Tables 2 shock depends on a small number of genes that function in assess- and 3). Most of these strains are much more sensitive to heat ing the metabolic health of the cell and/or regulate its growth in shock than the rest of the population; only a few were more a changing environment. resistant. We found that there was very little overlap between this set of genes and the much larger number of genes identified SYSTEMS BIOLOGY in other recent studies (5–8) as being induced as part of the heat- eat stress has been the subject of hundreds of studies, many Hof which have sought to understand the role of single genes fi or gene families in mediating the response to heat shock, whereas Signi cance others have attempted to examine system-level responses to heat (1, 2). Two basic approaches have been used to study the global We used the model eukaryote Saccharomyces cerevisiae to cellular response to heat or other stresses. One approach follows investigate which genes are important for survival of heat the response to temperature shift at a genomic scale using gene- stress. Previously, this question was addressed by examining expression microarrays to define the set of genes whose expres- which genes are turned on by mild heat stress; in this study, we sion is altered (3, 4). This approach necessarily limits the stress examined gene-deletion mutants for increased sensitivity to to temperatures at which the cells remain viable and express their lethal heat stress. This approach reveals that these two sets genes. The other approach uses single- or pooled-deletion strains of genes are largely nonoverlapping, demonstrating that mu- to assess the relative fitness of individuals alone or within a popu- tant analysis is a powerful complementary approach to gene- lation undergoing heat stress; this approach allows the use of lethal expression analysis. In addition, many of the genes identified heat to assess the ability to survive (5–8). as important for heat survival are involved in metabolism or We studied heat-stress resistance at a genomic scale further signaling, or their function is completely uncharacterized, sug- by introducing a number of experimental refinements. First, we gesting that our understanding of the systems-level response to used cultures growing at steady state before the heat stress to heat stress is incomplete. minimize effects of growth conditions. We had shown previously that the growth rate of cells has a profound effect on their ability Author contributions: P.A.G., C.L., A.A.C., D.C.H., and D.B. designed research; P.A.G., C.L., A.A.C., and D.C.H. performed research; P.A.G., C.L., A.A.C., and D.C.H. contributed new to survive a lethal heat shock (9): Slow-growing cells are much reagents/analytical tools; P.A.G., C.L., A.A.C., D.C.H., and D.B. analyzed data; and P.A.G., less sensitive than cells growing rapidly. By using the chemostat, C.L., A.A.C., D.C.H., and D.B. wrote the paper. variation in growth rate was minimized. Second, we used a newly The authors declare no conflict of interest. constructed prototrophic, barcoded nonessential deletion col- Freely available online through the PNAS open access option. lection to avoid the unwanted survival behaviors of auxotrophic 1P.A.G. and C.L. contributed equally to this work. mutations present in the standard, commercially available de- 2Present address: Banting and Best Department of Medical Research, Terrence Donnelly letion collection (7). Third, we used next-generation sequencing Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, ON, and our previously developed analysis algorithms to follow in- Canada M5S 3E1. dividual barcodes and thereby quantitatively assess relative sur- 3A.A.C. and D.C.H. contributed equally to this work. vival during heat stress (7, 10). 4To whom correspondence should be addressed. E-mail: [email protected]. Both thermotolerance and acquired thermotolerance have This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10. been studied in detail, with a general focus on the roles of transcripts 1073/pnas.1318100110/-/DCSupplemental. www.pnas.org/cgi/doi/10.1073/pnas.1318100110 PNAS | Published online October 28, 2013 | E4393–E4402 Downloaded by guest on October 1, 2021 Table 1. Phenotypic categories of heat-sensitive mutants 30–50 °C phenotype 37–50 °C phenotype Description Sensitive (P < 0.01) Sensitive (P < 0.01) Strains are hypersensitive to heat when shifted from 30–50 °C and do not acquire thermotolerance after pretreatment at 37 °C (10 genes; Table 2) Sensitive (P < 0.01) Resistant (P value for Strains are hypersensitive to heat when shifted from 30–50 °C and do acquire sensitivity is > 0.01) thermotolerance after pretreatment at 37 °C (49 genes; Table 3) Wild-type (P value for Sensitive (P < 0.05) Strains exhibit sensitivity to heat similar to wild type when shifted from 30–50 °C but exhibit sensitivity is > 0.05) hypersensitivity to lethal heat shock after pretreatment at 37 °C (nine genes; Table 4) shock response. Interestingly, many of the 55 genes we identified resource without this drawback, we generated a prototrophic as conferring hypersensitivity to heat are functionally involved deletion collection consisting of 4,783 viable deletion strains (see with cellular signaling, chromatin regulation, or carbohydrate Materials and Methods for details). This prototrophic deletion metabolism, suggesting unknown and potentially interesting collection has a number of useful features. First, each deletion is connections between heat shock and these processes. In the marked with the same barcode used in the original yeast deletion second experiment (pretreatment at 37 °C), we found only 10 collection. Thus, array-based methods for measuring deletion deletion strains that show significantly lower rates of survival frequency in pooled samples of the deletion collection will work than the pooled population as a whole (Table 2). These strains with this set. Second, the presence of the magic marker (can1Δ:: generally appear to be a subset of the 55 genes found in the first STE2pr-SpHIS5) allows the addition of another reporter (e.g., experiment, although the data for a few of them fell below our CLONAT-resistance) using the synthetic genetic array strategy (16, stringent threshold. We also found a third set of deletions (nine 17). Third, each deletion is marked with the G418-resistance cas- strains) whose survival in the first experiment (30–50 °C) was sette, as in the original deletion collection, allowing the user to select typical of the rest of the population but whose thermotolerance for G418-resistance when manipulating the deletion collection and was reduced significantly (by the same criteria used above) in the thus reducing potential contaminants. Fourth, unlike the original second experiment (Table 4). deletion collection, we included 12 isogenic wild-type controls on In this study we found only a few genes whose absence had a every 96-well plate as controls for position-related variance.
Load more

Recommended publications
  • Exploring Small-Scale Chemostats to Scale up Microbial Processes: 3-Hydroxypropionic Acid Production in S
    Lawrence Berkeley National Laboratory Recent Work Title Exploring small-scale chemostats to scale up microbial processes: 3-hydroxypropionic acid production in S. cerevisiae. Permalink https://escholarship.org/uc/item/43h8866j Journal Microbial cell factories, 18(1) ISSN 1475-2859 Authors Lis, Alicia V Schneider, Konstantin Weber, Jost et al. Publication Date 2019-03-11 DOI 10.1186/s12934-019-1101-5 Peer reviewed eScholarship.org Powered by the California Digital Library University of California Lis et al. Microb Cell Fact (2019) 18:50 https://doi.org/10.1186/s12934-019-1101-5 Microbial Cell Factories RESEARCH Open Access Exploring small-scale chemostats to scale up microbial processes: 3-hydroxypropionic acid production in S. cerevisiae Alicia V. Lis1, Konstantin Schneider1,2, Jost Weber1,3, Jay D. Keasling1,4,5,6,7, Michael Krogh Jensen1 and Tobias Klein1,2* Abstract Background: The physiological characterization of microorganisms provides valuable information for bioprocess development. Chemostat cultivations are a powerful tool for this purpose, as they allow defned changes to one single parameter at a time, which is most commonly the growth rate. The subsequent establishment of a steady state then permits constant variables enabling the acquisition of reproducible data sets for comparing microbial perfor- mance under diferent conditions. We performed physiological characterizations of a 3-hydroxypropionic acid (3-HP) producing Saccharomyces cerevisiae strain in a miniaturized and parallelized chemostat cultivation system. The physi- ological conditions under investigation were various growth rates controlled by diferent nutrient limitations (C, N, P). Based on the cultivation parameters obtained subsequent fed-batch cultivations were designed. Results: We report technical advancements of a small-scale chemostat cultivation system and its applicability for reliable strain screening under diferent physiological conditions, i.e.
    [Show full text]
  • Chemostat Culture for Yeast Experimental Evolution
    Downloaded from http://cshprotocols.cshlp.org/ at Cold Spring Harbor Laboratory Library on August 9, 2017 - Published by Cold Spring Harbor Laboratory Press Protocol Chemostat Culture for Yeast Experimental Evolution Celia Payen and Maitreya J. Dunham1 Department of Genome Sciences, University of Washington, Seattle, Washington 98195 Experimental evolution is one approach used to address a broad range of questions related to evolution and adaptation to strong selection pressures. Experimental evolution of diverse microbial and viral systems has routinely been used to study new traits and behaviors and also to dissect mechanisms of rapid evolution. This protocol describes the practical aspects of experimental evolution with yeast grown in chemostats, including the setup of the experiment and sampling methods as well as best laboratory and record-keeping practices. MATERIALS It is essential that you consult the appropriate Material Safety Data Sheets and your institution’s Environmental Health and Safety Office for proper handling of equipment and hazardous material used in this protocol. Reagents Defined minimal medium appropriate for the experiment For examples, see Protocol: Assembly of a Mini-Chemostat Array (Miller et al. 2015). Ethanol (95%) Glycerol (20% and 50%; sterile) Yeast strain of interest Equipment Agar plates (appropriate for chosen strain) Chemostat array Assemble the apparatus as described in Miller et al. (2013) and Protocol: Assembly of a Mini-Chemostat Array (Miller et al. 2015). Cryo deep-freeze labels Cryogenic vials Culture tubes Cytometer (BD Accuri C6) Glass beads, 4 mm (sterile; for plating yeast cells) Glass cylinder Kimwipes 1Correspondence: [email protected] © 2017 Cold Spring Harbor Laboratory Press Cite this protocol as Cold Spring Harb Protoc; doi:10.1101/pdb.prot089011 559 Downloaded from http://cshprotocols.cshlp.org/ at Cold Spring Harbor Laboratory Library on August 9, 2017 - Published by Cold Spring Harbor Laboratory Press C.
    [Show full text]
  • Evobot: an Open-Source, Modular, Liquid Handling Robot for Scientific Experiments
    applied sciences Article EvoBot: An Open-Source, Modular, Liquid Handling Robot for Scientific Experiments Andres Faiña 1,* , Brian Nejati 2 and Kasper Stoy 1 1 Robots, Evolution and Art Lab (REAL), Department of Computer Science, IT University of Copenhagen, 2300 Copenhagen, Denmark; [email protected] 2 Department of Mechanical and Industrial Engineering, University of Toronto, Toronto, ON M5S 1A1, Canada; His work was carried out while he was employed at ITU; [email protected] * Correspondence: [email protected]; Tel.: +45-7218-5258 Received: 13 October 2019; Accepted: 17 January 2020; Published: 23 January 2020 Featured Application: Our modular liquid handling robot can be applied to automate scientific experiments in research labs, where tasks change often, and to carry out new kinds of experiments that cannot be done manually. Abstract: Commercial liquid handling robots are rarely appropriate when tasks change often, which is the case in the early stages of biochemical research. In order to address it, we have developed EvoBot, a liquid handling robot, which is open-source and employs a modular design. The combination of an open-source and a modular design is particularly powerful because functionality is divided into modules with simple, well-defined interfaces, hence customisation of modules is possible without detailed knowledge of the entire system. Furthermore, the modular design allows end-users to only produce and assemble the modules that are relevant for their specific application. Hence, time and money are not wasted on functionality that is not needed. Finally, modules can easily be reused. In this paper, we describe the EvoBot modular design and through scientific experiments such as basic liquid handling, nurturing of microbial fuel cells, and droplet chemotaxis experiments document how functionality is increased one module at a time with a significant amount of reuse.
    [Show full text]
  • Raman Spectroscopy for the Microbiological Characterization and Identification of Medically Relevant Bacteria Khozima Mahmoud Hamasha Wayne State University
    Wayne State University Wayne State University Dissertations 1-1-2011 Raman spectroscopy for the microbiological characterization and identification of medically relevant bacteria Khozima Mahmoud Hamasha Wayne State University, Follow this and additional works at: http://digitalcommons.wayne.edu/oa_dissertations Part of the Physics Commons Recommended Citation Hamasha, Khozima Mahmoud, "Raman spectroscopy for the microbiological characterization and identification of medically relevant bacteria" (2011). Wayne State University Dissertations. Paper 311. This Open Access Dissertation is brought to you for free and open access by DigitalCommons@WayneState. It has been accepted for inclusion in Wayne State University Dissertations by an authorized administrator of DigitalCommons@WayneState. RAMAN SPECTROSCOPY FOR THE MICROBIOLOGICAL CHARACTERIZATION AND IDENTIFICATION OF MEDICALLY RELEVANT BACTERIA by KHOZIMA MAHMOUD HAMASHA DISSERTATION Submitted to the Graduate School of Wayne State University, Detroit, Michigan in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY 2011 MAJOR: PHYSICS Approved by: Advisor Date DEDICATION I would like to dedicate my dissertation to the memory of my brother “Saleh”. ii ACKNOWLEDGMENTS First of all, I would like to express my great thanks to my advisor Dr. Steven Rehse for his guidance and great help and support throughout my research work. I am always appreciative of this precious opportunity to learn from him about all research aspects, starting from thinking about the problems, sharing the ideas, performing the experiments, collecting the data, analyzing the results, and preparing the manuscripts for publication. Also, special thanks to the committee members, Dr. Ratna Naik for her continued support and encouragement through my graduate study, Dr. Choong-Min Kang for his great assistance and generosity through providing his facility to improve my skills and to getting trained in every microbiological aspect related to my research, and Dr.
    [Show full text]
  • Surface Growth of a Motile Bacterial Population Resembles Growth in a Chemostat
    Surface Growth of a Motile Bacterial Population Resembles Growth in a Chemostat Daniel A. Koster, Avraham Mayo, Anat Bren and Uri Alon Department of Molecular Cell Biology, Weizmann Institute of Science, Rehovot 76100, Israel Correspondence to Daniel A. Koster and Uri Alon: [email protected]; [email protected] http://dx.doi.org/10.1016/j.jmb.2012.09.005 Edited by I. B. Holland Abstract The growth behavior in well‐mixed bacterial cultures is relatively well understood. However, bacteria often grow in heterogeneous conditions on surfaces where their growth is dependent on spatial position, especially in the case of motile populations. For such populations, the relation between growth, motility and spatial position is unclear. We developed a microscope-based assay for quantifying in situ growth and gene expression in space and time, and we observe these parameters in populations of Escherichia coli swimming in galactose soft agar plates. We find that the bacterial density and the shape of the motile population, after an initial transient, are constant in time. By considering not only the advancing population but also the fraction that lags behind, we propose a growth model that relates spatial distribution, motility and growth rate. This model, that is similar to bacterial growth in a chemostat predicts that the fraction of the population lagging behind is inversely proportional to the velocity of the motile population. We test this prediction by modulating motility using inducible expression of the flagellar sigma factor FliA. Finally, we observe that bacteria in the chemotactic ring express higher relative levels of the chemotaxis and galactose metabolism genes fliC, fliL and galE than those that stay behind in the center of the plate.
    [Show full text]
  • Viscosity on Biofilm Phenotypic Expression
    University of Calgary PRISM: University of Calgary's Digital Repository Graduate Studies Legacy Theses 1999 Effect of viscosity on biofilm phenotypic expression Chan, Wendy Wing Yan Chan, W. W. (1999). Effect of viscosity on biofilm phenotypic expression (Unpublished master's thesis). University of Calgary, Calgary, AB. doi:10.11575/PRISM/12502 http://hdl.handle.net/1880/25429 master thesis University of Calgary graduate students retain copyright ownership and moral rights for their thesis. You may use this material in any way that is permitted by the Copyright Act or through licensing that has been assigned to the document. For uses that are not allowable under copyright legislation or licensing, you are required to seek permission. Downloaded from PRISM: https://prism.ucalgary.ca Wet of Viscosity on Biofilm Phenotypic Expression by Wendy Wing Yan Chan A THESIS SUBbWTO THE FACULTY OF GWUATE STUDIES IN PARTIAL FULFlLEiENT OF THE REQ~~TSFOR THE DEGREE OF bUSTER OF SCIENCE DEP-ARTMENT OF BIOLOGICAL SCIENCES CALGAELY? ALBERTA DECEMBm I999 OWendy Wing Yan Chan I999 National Library Bibliothe ue nationale du Cana1 a Acquisitions and Acquisitions et Bibliographic Services setvices bibliographiques 395 Wellington Street 395, rue Wellington Ortawa ON K1AON4 Ottawa ON KIA ON4 camdm Canada The author has granted a non- L'auteur a accorde me licence non exc1usive licence allowing the exclusive pennettant a la National h'brary of Canada to Bbliotheque nationale du Canada de reproduce, loan, distribute or sell reproduire, prster, distniuer ou copies of this thesis m microform, vendre des copies de cette these sous paper or electronic formats. la fome de microfiche/^ de reproduction sm papier ou sur format eIectroniqne.
    [Show full text]
  • Growth of an Adherent Mixed Microbial Culture in a Substrate
    WATER 171 PROJECT COMPLETION i REPORT NO. 391X Growth of an WATER Adherent Mixed Microbial Culture in a Substrate Limited WATER Single State Chemostat Robert M. Pfister Professor Department of Microbiology The Ohio State University WATER September, 1973 United States Department WATER of the Interior CONTRACT NO. WATER A-021-OHIO State of Ohio WATER Water Resources Center WATER Ohio State University GROWTH OF AN ADHERENT MIXED MICROBIAL CULTURE IN A SUBSTRATE LIMITED SINGLE STATE CHEMOSTAT by Robert M. Pfister Professor Department of Microbiology The Ohio State University Water Resources Center The Ohio State University Columbus, Ohio 43210 September 1973 This study was supported by the Office of Water Resources Research U.S. Department of the Interior under Project Number A- 021- OHIO TABLE OF CONTENTS page Summary • 1 Introduction 1 Methods 1 Results 6 Discussion 10 References 14 LIST OF FIGURES page 1* The single stage continuous culture apparatus (chemostat) 3 LIST OF TABLES Batch cultures of the Psdueomonas j3£, C. lividum and an equal mixture of the two microorganisms•. 7 2. Continuous cultivation of the Pseudomonas sp and °f £L* lividum in separate chemostats • 8 3. Continuous cultivation of the Psdueomonas in citrate medium 9 4. Continuous cultivation of the Psdueomonas sp in citrate medium followed by the inoculation of C. lividum 11 5. Continuous cultivation of C. lividum in citrate medium followed by the inoculation of Pseudomonas cells 12 SUMMARY A steady-state was established between a ^C. lividum and a Pseudomonas sp at a dilution rate of 0.27 hr when the growth limiting substrate was citrate, During both pure and mixed continuous culture studies, the jC.
    [Show full text]
  • Developing a Low-Cost Milliliter-Scale Chemostat Array for Precise Control of Cellular Growth
    Quantitative Biology 2018, 6(2): 129–141 https://doi.org/10.1007/s40484-018-0143-8 RESEARCH ARTICLE Developing a low-cost milliliter-scale chemostat array for precise control of cellular growth David Skelding1,*, Samuel F M Hart1, Thejas Vidyasagar2, Alexander E Pozhitkov1 and Wenying Shou1,* 1 Division of Basic Sciences, Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA 2 University of Washington, Seattle, WA 98195-3770, USA * Correspondence: [email protected], [email protected] Received November 24, 2017; Revised February 6, 2018; Accepted February 27, 2018 Background: Multiplexed milliliter-scale chemostats are useful for measuring cell physiology under various degrees of nutrient limitation and for carrying out evolution experiments. In each chemostat, fresh medium containing a growth rate-limiting metabolite is pumped into the culturing chamber at a constant rate, while culture effluent exits at an equal rate. Although such devices have been developed by various labs, key parameters — the accuracy, precision, and operational range of flow rate — are not explicitly characterized. Methods: Here we re-purpose a published multiplexed culturing device to develop a multiplexed milliliter-scale chemostat. Flow rates for eight chambers can be independently controlled to a wide range, corresponding to population doubling times of 3~13 h, without the use of expensive feedback systems. Results: Flow rates are precise, with the maximal coefficient of variation among eight chambers being less than 3%. Flow rates are accurate, with average flow rates being only slightly below targets, i.e., 3%‒6% for 13-h and 0.6%‒ 1.0% for 3-h doubling times. This deficit is largely due to evaporation and should be correctable.
    [Show full text]
  • Dunham Lab Chemostat Manual
    Dunham Lab Chemostat Manual Maitreya Dunham Last revised December 2005 This comprehensive manual covers the entire process of running a chemostat, including media recipes, chemostat setup, inoculation, data acquisition and storage, daily monitoring, harvests for DNA and RNA, and data analysis. Although I wrote it with my own yeast cultures and ATR Sixfors setup in mind, many of the procedures are general. If you have edits or additions to the manual, please email me at [email protected] Please feel free to point other people to these instructions. Also, I would appreciate the citation if you use any of this information in a publication or talk. Visit genomics.princeton.edu/dunham for the most recent updates to the protocol and for my other protocols on microarrays. Thanks to Matt Brauer, my long-time companion in the chemostat lab, for help developing these protocols and for proofing the manual. Also thanks to Frank Rosenzweig who taught me how to run my first glass-blown chemostats. Many of the protocols were influenced by his chemostat aesthetic. Edits since last version: new phosphate limitation recipe, new DNA prep, new filter-sterilization method of media preparation © Maitreya Dunham 2005 1 Contents CONTENTS ........................................................................................ 2 FIGURES ........................................................................................... 5 PLANNING THE EXPERIMENT............................................................ 6 Signup ................................................................................................6
    [Show full text]
  • Microbial Growth
    5 Microbial Growth The curved bacterium I Bacterial Cell Division 118 III Measuring Microbial Growth 128 Caulobacter has been a 5.1 Cell Growth and Binary Fission 118 5.9 Microscopic Counts 128 model for studying the cell 5.2 Fts Proteins and Cell Division 118 5.10 Viable Counts 129 division process, including 5.3 MreB and Determinants of Cell 5.11 Turbidimetric Methods 131 how shape-determining Morphology 120 IV Temperature and Microbial proteins such as crescentin 5.4 Peptidoglycan Synthesis and Cell Growth 132 Division 121 (shown here stained red) 5.12 Effect of Temperature on Growth 134 give cells their distinctive II Population Growth 123 5.13 Microbial Life in the Cold 134 shape. 5.5 The Concept of Exponential 5.14 Microbial Life at High Growth 123 Temperatures 138 5.6 The Mathematics of Exponential V Other Environmental Factors Growth 124 5.7 The Microbial Growth Cycle 125 Affecting Growth 140 5.8 Continuous Culture: The 5.15 Acidity and Alkalinity 140 Chemostat 126 5.16 Osmotic Effects 141 5.17 Oxygen and Microorganisms 143 5.18 Toxic Forms of Oxygen 146 118 UNIT 2 • Metabolism and Growth I Bacterial Cell Division n the last two chapters we discussed cell structure and function Cell I(Chapter 3) and the principles of microbial nutrition and elongation metabolism (Chapter 4). Before we begin our study of the biosyn- thesis of macromolecules in microorganisms (Chapters 6 and 7), we consider microbial growth. Growth is the ultimate process in the life of a cell—one cell becoming two. Septum Septum formation 5.1 Cell Growth and Binary Fission In microbiology, growth is defined as an increase in the number One generation Completion of of cells.
    [Show full text]
  • A High-Throughput Platform for Feedback-Controlled Directed Evolution
    bioRxiv preprint doi: https://doi.org/10.1101/2020.04.01.021022; this version posted April 2, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. A high-throughput platform for feedback-controlled directed evolution Erika A. DeBenedictis1,2, Emma J. Chory1,2*, Dana Gretton2*, Brian Wang2, Kevin Esvelt2 1Department of Biological Engineering, Massachusetts Institute of Technology. 2Department of Media Arts and Sciences, Massachusetts Institute of Technology. *Designates equal-contribution Continuous directed evolution rapidly implements cycles of mutagenesis, selection, and replication to accelerate protein engineering. However, individual experiments are typically cumbersome, reagent-intensive, and require manual readjustment, limiting the number of evolutionary trajectories that can be explored. We report the design and validation of Phage-and-Robotics-Assisted Near-Continuous Evolution (PRANCE), an automation platform for the continuous directed evolution of biomolecules that enables real-time activity- dependent reporter and absorbance monitoring of up to 96 parallel evolution experiments. We use this platform to characterize the evolutionary stochasticity of T7 RNA polymerase evolution, conserve precious reagents with miniaturized evolution volumes during evolution of aminoacyl-tRNA synthetases, and perform a massively parallel evolution of diverse candidate quadruplet tRNAs. Finally, we implement a feedback control system
    [Show full text]
  • 25% Off List Price
    SCIENTIFIC LABORATORY SUPPLIES PRESENTS F CUSEXCLUSIVE OFFERS & EXCITING NEW PRODUCTS Eppendorf 5430 & 5430R Centrifuge Packages NEW s NEW Integra Biosciences ee pa EVOLVE Manual Pipettes ge 11 25% off list price 6 ge see pa Brought to you by the UK’s largest independent supplier of scientific products May-Aug 2016 Welcome to the Scientific Laboratory Show FOCUS 2016 This edition of Focus brings you more new products and exclusive offers than ever before in conjunction with the Scientific Laboratory Show 2016. This year is set to be the biggest and best show yet, with all the offers, products and brands you see in this Focus on display at the event on Wednesday 25th May 2016. Whether your discipline is Life Sciences or Chemistry, Clinical or Analytical we have the product range to suit your needs and budget, underpinned by our ongoing commitment as the UK’s largest independent supplier of laboratory equipment. With an array of high profile Key Speakers presenting the Conference Programme, the Scientific Laboratory Show 2016 along with Key Supply Partners Eppendorf & Corning, provides delegates with an exciting occasion of real scientific value. Established as an integral part of the UK scientific calendar attracting delegates from over 20 countries, the event gives you an opportunity to see and discuss the latest products and developments from over 60 of the world’s leading manufacturers of laboratory products - all under one roof! At the NEW Professional Development Zone hosted by the Science Council you can take advantage of the opportunities to develop your career and professional experience. The Science Council has brought together experts, exhibitors and organisations providing one to one advice such as CV surgeries, 15 minute masterclasses on areas of professional development and membership offers exclusive to SLS Lab Show delegates.
    [Show full text]