PCR TROUBLESHOOTING

and OPTIMIZATION OOK !!!

The Essential Guide

Edited by: Suzanne Kennedy and Nick Oswald

240 pp., January 2011

ISBN 978-1-904455-72-1
Published by: Caister Academic Press

EW

$310/159
www.caister.com

The polymerase chain reaction (PCR) is a fundamental tool in scientific research and clinical
testing. Real-time PCR, combining both amplification and detection in one instrument, is a rapid
and accurate method for nucleic acid detection and quantification. Although PCR is a very powerful
technique, the results achieved are valid only if the appropriate controls have been employed.
In addition, proper optimization of PCR conditions is required for the generation of specific,
repeatable, reproducible and sensitive data.
This book discusses the strategies for preparing effective controls and standards for PCR,
when they should be employed and how to interpret the information they provide. It highlights
the significance of optimization for efficiency, precision and sensitivity of PCR methodology
and provides essential guidance on how to troubleshoot inefficient reactions. Experts in PCR
describe design and optimization techniques, discuss the use of appropriate controls, explain the
significance of standard curves and explore the principles and strategies required for effective
troubleshooting. Authors highlight the importance of sample preparation and quality, primer design,
controlling inhibitors, avoiding amplicon and environmental contamination, optimizing reagent
quality and concentration, and modifying the thermal cycling protocol for optimal sensitivity and
specificity. In addition, specific chapters discuss the history of PCR, the choice of instrumentation,
the applications of PCR in metagenomics, high resolution melting analysis, the MIQE guidelines,
and PCR at the microliter scale.
The strategies, tips and advice contained in this concise volume enable the scientist to optimize
and effectively troubleshoot a wide range of techniques including PCR, reverse transcriptase PCR,
real-time PCR and quantitative PCR. An essential book for anyone using PCR technology.

Table of Contents
Magic in Solution: An Introduction and
Brief History of PCR
Difficult Templates and Inhibitors of
PCR
Significance of Controls and Standard
Curves in PCR
Obtaining Maximum PCR Sensitivity
and Specificity
RT-PCR Optimization Strategies
Real-Time PCR Instrumentation: An
Instrument Selection Guide
qPCR Data Analysis: Unlocking the
Secret to Successful Results
The MIQE Guidelines Uncloaked
PCR Applications for Epigenetics
Research
High Resolution Melting Analysis
Microfluidic Emulsion PCR

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