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A-Z of Quantitative PCR

lots of recipes from distinguished contributors

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ISBN-10:
0-96368178-8
ISBN-13:
978-0-96368178-2

publication info

S. Bustin, ed., IUL Press, 2004, 830 pp., hard cover

technique(s)

PCR: suitable

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General description

This is not just a cookbook for real-time quantitative PCR (qPCR). Admittedly, there are lots of recipes from distinguished contributors and Bustin has attempted to collect, sift through and rationalize the vast amount of information that is available on this subject. And yes, this book was conceived as a comprehensive hands-on manual to allow both the novice researcher and the expert to set up and carry out qPCR assays from scratch.

However, this book also sets out to explain as many features of qPCR as possible, provide alternative viewpoints and methods and, perhaps most importantly, aims to stimulate the researcher into generating, interpreting and publishing data that are reproducible, reliable and biologically meaningful.

Table of Contents

PART I REVIEWS
1. Quantification of nucleic acids by PCR
2. Real-time RT-PCR: what lies beneath the surface?
3. Quantification strategies in real-time PCR

PART II
REAL-TIME PCR - THE BASICS
1. Good laboratory practice
2. Sample acquisition, template preparation, quantification and quality assessment
3. Principles of fluorescence and real-time chemistries
4. Probes and primers
5. Principles of real-time detection and instrumentation
6. Basic RT-PCR considerations
7. The PCR step
8. Data Analysis and interpretation
9. Troubleshooting

PART III
PROTOCOLS
1. Getting started
2. Use of standardized mixtures of internal standards in quantitative RT-PCR to ensure quality control and develop a standardized gene expression database
3. Standardization of qPCR and qRT-PCR assays
4. Extraction of RNA from formalin-fixed paraffin-embedded archival material
5. Cells-to-cDNA II: RT-PCR without RNA Isolation
6. Optimization of Single and Multiplex Real-Time PCR
7. Evaluation of Basic Fibroblast Growth FActor mRNA levels in Breast Cancer
8. Detection of cytokeratin 20 mRNA in the blood and lymph nodes of patients without colorectal cancer
9. Optimized Real-time RT-PCR for Quantitative Measurements of DNA and RNA in Single Embryos and Blastomeres
10. Single cell global RT and quantitative real-time PCR
11. Single nucleotide polymorphism detection with fluorescent minor groove binder probes
12. Genotyping using MGB-hydrolysis probes
13. Scorpion primers for real time genotyoping and quantitative genotyping on pooled DNA
14. Simultaneous Detection and Sub-typing of Human Papillomavirus in the Cervix Using Real-time Quantitative PCR

PART IV
APPENDIX
Conversions and calculations
Glossary

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Anja Hemschemeier et al.
Eukaryotic cell, 7(3), 518-526 (2008-02-05)
The unicellular green alga Chlamydomonas reinhardtii has a special type of anaerobic metabolism that is quite unusual for eukaryotes. It has two oxygen-sensitive [Fe-Fe] hydrogenases (EC 1.12.7.2) that are coupled to photosynthesis and, in addition, a formate- and ethanol-producing fermentative

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