Definitive qPCR: A practical guide

Definitive qPCR: A practical guide

Revised February 2015

This volume deals with the practical steps required to characterise, optimise and validate a qPCR assay. It deals with all aspects of laboratory organisation, instruments, pipettes, plasticware and reagents as well as providing protocols for optimisation of annealing temperatures, primer concentration and determining assay efficiency. The 320 pages are richly illustrated with examples, tips, explanations and all information is hyperlinked to provide access to the huge amount of information that is available online, but not always found in obvious places.

This book will be invaluable to anyone wanting to design, validate or trouble shoot qPCR assays. Its easily accessible language takes the reader through the complex steps that make up a successful qPCR assay and will result, it is hoped, in data that are accurate, robust and biologically relevant.

Contents
1. The Basics
Why optimise and validate
Laboratory environment
Thermal cyclers
Plasticware
Pipettes
Reagents
Oligonucleotide handling
2. Annealing temperature
Optimisation of Ta
3. Primer Concentration
Optimisation of primers
4. PCR efficiency
Assay optimisation
5. MIQE
Getting it wrong in the laboratory
6. Appendices
Definitive qPCR: A practical guide
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