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E-raamat: Pyrosequencing: Methods and Protocols

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The primary purpose of this volume is to demonstrate the range of applications of the Pyrosequencing technology in research and diagnostics and to provide detailed protocols. Beginning with an up-to-date overview of the biochemistry, the volume continues with quantitative analysis of genetic variation, ratio of expressed alleles at the RNA level, analysis of DNA methylation, global DNA methylation assays, specialized applications for DNA methylation analysis including loss of imprinting, single blastocyst analysis, allele-specific DNA methylation patterns, DNA methylation patterns associated with specific histone modifications. The volume further details tools and protocols for the detection of viruses and bacteria, and genetic and epigenetic analyses for forensics using Pyrosequencing. As a volume in the highly successful Methods in Molecular Biology series, chapters contain introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible protocols and tips on troubleshooting and avoiding known pitfalls.

Comprehensive and adaptable, Pyrosequencing: Methods and Protocols, Second Edition will greatly aid doctorial students, postdoctoral investigators, and research scientists studying different aspects of genetics and cellular and molecular biology.

Preface v
Contributors xi
Part I Introduction
1 The History of Pyrosequencing®
3(14)
Pal Nyren
2 PyroMark® Instruments, Chemistry, and Software for Pyrosequencing® Analysis
17(12)
Martin Kreutz
Gerald Schock
Julia Kaiser
Norbert Hochstein
Ralf Peist
3 Software-Based Pyrogram® Evaluation
29(10)
Guoli Chen
Matthew T. Olson
James R. Eshleman
4 Quantitative Validation and Quality Control of Pyrosequencing® Assays
39(10)
Ulrich Lehmann
Part II Analysis of Genetic Variation
5 Extended KRAS and NRAS Mutation Profiling by Pyrosequencing®
49(14)
Andreas Jung
6 Universal BRAF State Detection by the Pyrosequencing®-Based U-BRAFV600 Assay
63(20)
Alexander Skorokhod
7 Pyrosequencing®-Based Identification of Low-Frequency Mutations Enriched Through Enhanced- w-COLD-PCR
83(20)
Alexandre How-Kit
Jorg Tost
8 Analysis of Mutational Hotspots in Routinely Processed Bone Marrow Trephines by Pyrosequencing®
103(12)
Stephan Bartels
Ulrich Lehmann
9 Analysis of Copy Number Variation by Pyrosequencing® Using Paralogous Sequences
115(8)
Marianne Kristiansen Kringen
10 Prenatal Diagnosis of Chromosomal Aneuploidies by Quantitative Pyrosequencing®
123(10)
Hut Ye
Haiping Wu
Yunlong Liu
Bingjie Zou
Tomoharu Kajiyama
Hideki Kambara
Guohua Zhou
11 HLA-B and HLA-C Supratyping by Pyrosequencing®
133(20)
Irene Vanni
Elisabetta Ugolotti
Patrizia Larghero
Roberto Biassoni
12 Allele Quantification Pyrosequencing® at Designated SNP Sites to Detect Allelic Expression Imbalance and Loss-of-Heterozygosity
153(22)
Chau-To Kwok
Megan P. Hitchins
Part III Analysis of DNA Methylation Patterns
13 Quantitative DNA Methylation Analysis by Pyrosequencing®
175(14)
Jessica Roessler
Ulrich Lehmann
14 Quantitative Methylation Analysis of the PCDHB Gene Cluster
189(12)
Barbara Banelli
Massimo Romani
15 Assessment of Changes in Global DNA Methylation Levels by Pyrosequencing® of Repetitive Elements
201(8)
Ali M. Tabish
Andrea A. Baccarelli
Lode Godderis
Timothy M. Barrow
Peter Hoet
Hyang-Min Byun
16 Global Analysis of DNA 5-Methylcytosine Using the Luminometric Methylation Assay, LUMA
209(12)
Karin Luttropp
Louise K. Sjoholm
Tomas J. Ekstrom
17 Limiting Dilution Bisulfite Pyrosequencing®: A Method for Methylation Analysis of Individual DNA Molecules in a Single or a Few Cells
221(20)
Nady El Hajj
Juliane Kuhtz
Thomas Haaf
18 Detection of Loss of Imprinting by Pyrosequencing®
241(18)
Silvia Tabano
Eleonora Bonaparte
Monica Miozzo
19 Analysis of DNA Methylation Patterns in Single Blastocysts by Pyrosequencing®
259(12)
John Huntriss
Kathryn Woodfine
Joanna E. Huddleston
Adele Murrell
Helen M. Picton
20 Allele-Specific DNA Methylation Detection by Pyrosequencing®
271(20)
Lasse Sommer Kristensen
Jens Vilstrup Johansen
Kirsten Grønbak
21 SNP-based Quantification of Allele-specific DNA Methylation Patterns by Pyrosequencing®
291(24)
Florence Busato
Jorg Tost
22 DNA Methylation Analysis of ChIP Products at Single Nucleotide Resolution by Pyrosequencing®
315(22)
Celine Moison
Fanny Assemat
Antoine Daunay
Paola B. Arimondo
Jorg Tost
Part IV Bacterial Typing and Identification
23 Multiplex Pyrosequencing®: Simultaneous Genotyping Based on SNPs from Distant Genomic Regions
337(12)
Piotr Wojciech Dabrowski
Kati Bourquain
Andreas Nitsche
24 Detection of Drug-Resistant Mycobacterium tuberculosis
349(14)
Anna Engstrom
Pontus Jureen
25 Application of Pyrosequencing® in Food Biodefense
363(16)
Kingsley Kwaku Amoako
Part V Forensics
26 Forensic Analysis of Mitochondrial and Autosomal Markers Using Pyrosequencing®
379(18)
Magdalena M. Bus
Hanna Edlund
Marie Allen
27 Tissue-Specific DNA Methylation Patterns in Forensic Samples Detected by Pyrosequencing®
397(14)
Joana Antunes
Kuppareddi Balamurugan
George Duncan
Bruce McCord
Index 411