Paper No. : 16 Module : 21 Methods for gene identification: In-situ Hybridization

Development Team

Principal Investigator: Prof. Neeta Sehgal Head, Department of Zoology, University of Delhi

Co-Principal Investigator: Prof. D.K. Singh Department of Zoology, University of Delhi

Paper Coordinator: Prof. Namita Agrawal Department of Zoology, University of Delhi

Content Writer: Dr. Jasvinder Kaur, Dr. Poonam Sharma, Gargi College, University of Delhi

Content Reviewer: Dr. Surajit Sarkar Department of Genetics, South Campus, University of Delhi

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ZOOLOGY Molecular Genetics In-situ Hybridization

Description of Module

Subject Name ZOOLOGY

Paper Name Zool 016: Molecular Genetics

Module Name/Title Methods for gene identification

Module ID M21: In-situ Hybridization

Keywords In-situ Hybridization, Probe, Radioactive label, Fish, Chromosome painting

Glossary

Avidin: Biotin binding protein, found in the egg whites of birds, amphibians and reptiles (produced in their oviducts).

Chromosome mapping: Technique using which genes belonging to same gene families can be detected by hybridization with suitable labeled probes.

Chromosome painting: Multiple fluorochromes can be used in FISH technique which produces a multicolored and painted effect with unique color at each hybridization site in the chromosomes being studied giving the effect of chromosome painting.

Fluorescent in-situ hybridization:It is a cytogenetic technique that uses fluorescent probes to detect the presence or absence of specific DNA sequences on chromosomes.

Hybridization probe:Probe is a fragment of DNA or RNA of variable length, which can be radioactively labelled. Used to detect sequence, complementary to the sequence in the probe.

Immunohistochemistry: Technique used to localize proteins such as antibodies in tissue sections by antigen-antibody interactions.

In-situ hybridization (ISH): ISH is a technique used to detect and localize specific mRNA sequences (within preserved tissues) and cells (within a heterogeneous cell population).It is based on the complementary base pairing of labelled cDNA, or, RNA probe to normal or abnormal sequences in chromosomes, cells or tissues.

Multiple ISH: This technique uses more than one probe to detect different nucleic acid targets in the same tissue section.

Oligonucleotide: Short DNA or RNA molecules having wide range of applications.

Polymerase chain reaction ISH: This is a variable ISH technique used for detection of targets present even in low copy numbers. 2

ZOOLOGY Molecular Genetics In-situ Hybridization

Probes: Complementary sequence of nucleotide bases to the specific sequence of interest such as mRNA are referred to as probes.

Riboprobes: RNA probes, 50-1000 bp in length. RNA-RNA hybrid probes are very thermostable and are resistant to digestion by RNases.

Q-FISH: This technique uses FISH and computer software to quantify fluorescence intensity.

References

1. Meng, L., and Leng, P. In situ hybridisation: principles and applications. 1992. Malaysian J Pathol. 14(2): 69 – 76. 2. Clare O'Connor. Fluorescence (FISH). 2008. Nature Education 1(1):171 Weblinks: http://www.powershow.com/view/1b405eNzBkN/In_Situ_Hybridization_powerpoint_pp t_presentation 3. http://www.genedetect.com/insitu.htm 4. https://en.wikipedia.org/wiki/In_situ_hybridization 5. http://www.ncbi.nlm.nih.gov/probe/docs/techish/

Further Readings:

1. Fluorescence In Situ Hybridization (FISH) - Application Guide. Edited by Thomas Liehr 2. In Situ Hybridization: Principles and Practice.Julia M. Polak, James O'Donnell McGee.Oxford University Press, 1998 3. In Situ Hybridization: A Practical Guide.A. R. Leitch.BIOS Scientific Publishers, 1994.

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ZOOLOGY Molecular Genetics In-situ Hybridization