PCR 3: PCR In Situ Hybridization A Practical Approach

by Herrington, C. Simon; O'Leary, John J.

• ISBN13: 9780199636327
• ISBN10: 019963632X
• Format: Paperback
• Copyright: 1998-01-08
• Publisher: Oxford University Press

Summary

PCR in situ hybridization allows the detection of specific nucleic acid sequences and their distribution in the cell. It combines two powerful techniquesin situ hybridization (ISH), which allows cellular localization of DNA and RNA sequences in cells and tissues, and the polymerase chainreaction (PCR), which allows reproducible amplification of rare nucleic acid sequences. The combined technique and its variants greatly enhance the sensitivity of in situ hybridization and add morphological localization to the sensitivity of PCR. Such techniques have enormous potential for researchand diagnosis but problems with reproducibility and reliability are often encountered. This book overcomes these problems by describing the key procedures in step-by-step detail and by providing the essential advice needed for success. Topics include: DNA in situ PCR and DNA PCR in situhybridization (PCR ISH) for the detection of DNA targets in cells; reverse trancriptase in situ PCR (RT-PCR) and RT-PCR ISH for the detection of RNA targets; and PRINS (primed in situ synthesis) for chromosomal analysis in interphase nuclei and metaphase chromosome spreads. There are furtherchapters on fixation of tissues for PCR, selective ultraviolet radiation fractionation (SURF), application of in situ PCR to human tissues, applications and modifications of PCR-ISH, and automation of in situ amplification. PCR In Situ Hybridization is a unique and timely collection of well-testedprotocols for the amplification of DNA and RNA in cells and tissues, drawing on the accumulated knowledge and experience of leading exponents of these techniques. For each topic covered, the authors provide detailed guidance on the key steps in the protocols, numerous hints and tips for success,and advice on trouble-shooting. PCR In Situ Hybridization will be invaluable to molecular biologists, pathologists, geneticists, and all those seeking to perform in situ analyses of nucleic acid molecules.

Table of Contents

Fixation of tissues for the polymerase chain reaction

In situ genetic analysis with selective ultraviolet radiation fractionation (SURF)

In situ hybridization

PCR in situ hybridization (PCR-ISH)

Reverse transcriptase in situ PCR for RNA detection

Primed in situ DNA synthesis (PRINS)

Application of in situ-PCR techniques to human tissues

Applications and modifications of PCR in situ hybridization

Automation of in situ amplification

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