Pyrosequencing (2^nd Ed., 2nd ed. 2015) Methods and Protocols Methods in Molecular Biology Series, Vol. 1315

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.

Pål Nyrén

2. PyroMark® Instruments, Chemistry, and Software for Pyrosequencing® Analysis

Martin Kreutz, Gerald Schock, Julia Kaiser, Norbert Hochstein, and Ralf Peist

3. Software-Based Pyrogram^® Evaluation

Guoli Chen, Matthew T. Olson, and James R. Eshleman

4. Quantitative Validation and Quality Control of Pyrosequencing^® Assays

Ulrich Lehmann

Part II: Analysis of Genetic Variation

5. Extended KRAS and NRAS Mutation Profiling by Pyrosequencing®

Andreas Jung

6. Universal BRAF State Detection by the Pyrosequencing®-based U‑BRAF^V600 Assay

Alexander Skorokhod

7. Pyrosequencing^®-based Identification of Low Frequency Mutations Enriched through Enhanced-ice-COLD-PCR

Alexandre How-Kit and Jörg Tos

8. Analysis of Mutational Hotspots in Routinely Processed Bone Marrow Trephines by Pyrosequencing^®

Stephan Bartels and  Ulrich Lehmann

9. Analysis of Copy Number Variation By Pyrosequencing^® using Paralogous Sequences

Marianne Kristiansen Kringen

10. Prenatal Diagnosis of Chromosomal Aneuploidies by Quantitative Pyrosequencing^®

Hui Ye, Haiping Wu, Yunlong Liu, Bingjie Zou, Tomoharu Kajiyama,  Hideki Kambara,and Guohua Zhou

11. HLA-B and HLA-C Supratyping by Pyrosequencing^®

Irene Vanni, Elisabetta Ugolotti, Patrizia Larghero, and Roberto Biassoni

12. Allele Quantification Pyrosequencing® at Designated SNP sites to Detect Allelic Expression Imbalance and Loss-of-heterozygosity

Chau-To Kwokand Megan P. Hitchins

Part III: Analysis of DNA Methylation Patterns

13. Quantitative DNA Methylation Analysis by Pyrosequencing®

14. ­Quantitative Methylation Analysis of the PCDHB Gene Cluster

Barbara Banelli and Massimo Romani

15. Assessment of Changes in global DNA Methylation Levels by Pyrosequencing® of Repetitive Elements

Ali M. Tabish, Andrea A. Baccarelli, Lode Godderis, Timothy M. Barrow, Peter Hoet¹ and Hyang-Min Byun

16. Global Analysis of DNA 5-methylcytosine using the Luminometric

Methylation Assay, LUMA

Karin Luttropp, Louise K Sjöholm, and Tomas J Ekström

17. Limiting Dilution Bisulfite Pyrosequencing^®: a Method for Methylation Analysis of Individual DNA Molecules in a Single or a Few Cells

Nady El Hajj, Juliane Kuhtz, and Thomas Haaf

18. Detection of Loss of Imprinting by Pyrosequencing^®

Silvia Tabano, Eleonora Bonaparte,  and Monica Miozzo

19. Analysis of DNA Methylation Patterns in Single Blastocysts by Pyrosequencing^®

John Huntriss, Kathryn Woodfine, Joanna E Huddleston, Adele Murrell, and  Helen Mary Picton

20. Allele-specific DNA methylation detection by Pyrosequencing^®

Lasse Sommer Kristensen, Jens Vilstrup Johansen, and Kirsten Grønbæk

21. SNP-based Quantification of Allele-Specific DNA Methylation Patterns by Pyrosequencing^®

Florence Busatoand Jörg Tost

22. DNA Methylation Analysis of ChIP Products at Single Nucleotide Resolution byPyrosequencing^®

Céline Moison, Fanny Assemat, Antoine Daunay, Paola B Arimondo, and Jörg Tost

Part IV: Bacterial Typing and Identification

23. Multiplex Pyrosequencing®: Simultaneous Genotyping Based on SNPs from Distant Genomic Regions

Piotr Wojciech Dabrowski, Kati Bourquain, and Andreas Nitsche

24. Detection of Drug-Resistant Mycobacterium tuberculosis

Anna Engströmand Pontus Juréen

25. Application of Pyrosequencing^® in Food Biodefense

Kingsley Kwaku Amoako

Part V: Forensics

26. Forensic Analysis of Mitochondrial and Autosomal Markers using Pyrosequencing^®

Magdalena M. Buś, Hanna Edlund, and Marie Allen

27. Tissue-specific DNA Methylation Patterns in Forensic Samples Detected By Pyrosequencing^®

Joana Antunes, Kuppareddi Balamurugan, George Duncan, and Bruce McCord

Includes cutting-edge methods and protocols

Provides step-by-step detail essential for reproducible results

Contains key notes and implementation advice from the experts