Next Generation Sequencing in Forensic Science: A Primer

Description

Next Generation Sequencing in Forensic Science: A Primer addresses next generation sequencing (NGS) specific to its application to forensic science. The first part of the book offers a history of human identity approaches, including VNTR, RFLP, STR, and SNP DNA typing. It discusses the history of sequencing for human DNA typing, including Sanger sequencing, SNaPshot, pyrosequencing, and principles of next generation sequencing. The chapters present an overview of the forensically focused AmpliSeq, ForenSeq, Precision ID, PowerSeq, and QIAseq panels for human DNA typing using autosomal, Y and X chromosome STRs and SNPs using the MiSeq FGx and Ion Torrent System. The authors outline the steps included in DNA extraction and DNA quantitation that are performed prior to preparing libraries with the NGS kits. The second half of the book details the implementation of ForenSeq and Precision ID to amplify and tag targets to create the library, enrich targets to attach indexes and adaptors, perform library purification and normalization, pool the libraries, and load samples to the cartridge to perform the sequencing on the instrument. Coverage addresses the operation of the MiSeq FGx and Ion Chef, including creating a sample list, executing wash steps, performing NGS, understanding the run feedback files from the instrument, and troubleshooting. ForenSeq and Precision ID panel data analysis are explained, including how to analyze and interpret NGS data and output graphs and charts. The book concludes with mitochondrial DNA (mtDNA) sequencing and SNPs analysis, including the issue of heteroplasmy. The final chapters review forensic applications of microbial DNA, NGS in body fluid analysis, and challenges and considerations for future applications. FEATURES Focuses on human identification using traditional and NGS DNA typing methods targeting short tandem repeats (STRs) Applies the technology and its application to law enforcement investigations and identity and ancestry single nucleotide polymorphisms (SNPs) for investigational leads, mass disaster, and ancestry cases Presents the underlying principles of NGS in a clear, easy-to-understand format for practitioners and students studying DNA in forensic programs This is the first book to prepare practitioners to utilize and implement this new technology in their lab for casework, highlighting early applications of how NGS results have been used in court. The book can be utilized for upper-level undergraduate and graduate students taking courses focused on NGS concepts. Readers are expected to have a basic understanding of molecular and cellular biology and DNA typing. Cover Half Title Title Page Copyright Page Dedication Table of Contents Foreword Preface Acknowledgments Authors List of Figures List of Tables List of Credits List of Abbreviations 1 History of DNA-Based Human Identification in Forensic Science 1.1 Introduction 1.2 Application of DNA Sequencing to Human DNA 1.3 History of DNA Typing 1.4 Next Generation Sequencing for Forensic DNA Typing 1.5 Conclusion Questions References 2 History of Sequencing for Human DNA Typing 2.1 Introduction 2.2 Common Chemistries Used in Sequencing Applications 2.2.1 Chain Termination Sequencing 2.2.2 Pyrosequencing 2.2.3 Sequencing by Ligation 2.3 Detection Techniques 2.3.1 Fluorescence 2.3.2 Pyrosequencing 2.3.3 Ion Detection 2.4 Sequencing Platforms 2.4.1 First-Generation Sequencing Techniques 2.4.1.1 Sanger Sequencing 2.4.1.2 SNaPShot Sequencing 2.4.1.3 Pyrosequencing 2.5 Massively Parallel Sequencing 2.5.1 Reversible Chain Termination MPS Platforms 2.5.2 Ion Detection Platforms 2.5.3 Sequencing by Ligation Platforms 2.5.4 Single Base Extension Platforms 2.5.5 Third-Generation Platforms 2.6 NGS Instruments Adopted for Forensic Science Questions References 3 Sample Preparation, Standards, and Library Preparation for Next Generation Sequencing 3.1 Overview of the NGS Sample Preparation Process 3.2 Sample Handling and Processing 3.3 DNA Extraction 3.4 DNA Quantitation 3.5 Library Preparation 3.6 Library Purification and Normalization 3.7 Multiplexing and Denaturation Questions References 4 Performing Next Generation Sequencing 4.1 Performing Next Generation Sequencing 4.2 Verogen MiSeq FGx[sup(®)] Sequencing 4.3 ThermoFisher Ion Torrent[sup(™)] and Ion PGM Sequencing 4.4 The Next Step Questions References 5 Next Generation Sequencing Data Analysis and Interpretation 5.1 NGS Data Analysis 5.2 Verogen Universal Analysis Software 5.3 ThermoFisher Converge Software 5.4 Phenotype Analysis Using the Erasmus Server 5.5 Other Sequence Analysis Software 5.6 Additional Tools for Mixture Interpretation 5.7 Other NGS Sequence Data Analysis Tools 5.8 NGS Validation and Applications Questions References 6 Next Generation Sequencing Troubleshooting 6.1 Troubleshooting NGS Sequencing 6.2 Troubleshooting MiSeq FGx Instrument Failure 6.3 Troubleshooting MiSeq FGx Run Failure 6.4 Troubleshooting Ion Series Run Failure Questions References 7 Mitochondrial DNA Typing Using Next Generation Sequencing 7.1 Introduction to Mitochondrial DNA Typing 7.2 The Sequence of the Mitochondrial Chromosome 7.3 Mitochondrial DNA Typing Methods 7.4 Mitochondrial DNA Typing Using Next Generation Sequencing 7.5 Mitochondrial Sequence Data Interpretation and Reporting 7.6 Recent Reports of Mitotyping Using NGS for Forensic Applications 7.7 Mitochondrial Sequence Data and Databases Questions References 8 Microbial Applications of Next Generation Sequencing for Forensic Investigations 8.1 Introduction to Microbial DNA Profiling 8.2 Why NGS? 8.3 The Human Microbiome Project 8.4 Sampling and Processing 8.5 NGS Methodology in Microbial Forensics 8.6 Results from the Human Microbiome Project 8.7 HMP Applications for Forensic Science 8.8 NGS Applications in Geolocation, Autopsy, PMI, and Lifestyle Analysis 8.9 Bioinformatic Approaches and Tools 8.10 Bioforensics and Biosurveillance 8.11 Infectious Disease Diagnostics 8.12 NGS Applications in Archeology 8.13 Summary of NGS Microbial Sequencing Applications in Forensic Investigation Questions References 9 Body Fluid Analysis Using Next Generation Sequencing 9.1 Introduction 9.2 Epigenetic-Based Tissue Source Attribution 9.3 mRNA-Based Tissue Source Attribution 9.4 MicroRNA Analysis 9.5 The Future of Body Fluid Assays Questions References 10 Conclusions and Future Outlook of Next Generation Sequencing in Forensic Science 10.1 NGS Is Here 10.2 Why NGS? 10.3 Ongoing Challenges of Adopting NGS for Forensic Investigations 10.4 Early Successes of NGS in Forensic Cases 10.5 Summary Questions References Index