Molecular Microbiology: Diagnostic Principles and Practice, 2 nd edition

Abstract

Over the last decade, next-generation sequencing (NGS) became one of the fastest-growing biotechnologies. Rapid advances in sequencing technology, library preparation, and data analysis tools have driven sequencing costs down and have contributed to the emergence of many diverse applications across many different areas. Furthermore, the introduction of several small, personal sequencing devices have contributed to the democratization of the sequencing field. Complex sequencing projects that until recently could have been carried out only in major sequencing centers can now be performed by medium-sized research groups or small research facilities. In this chapter, we present an introductory overview of the NGS field, with the emphasis on various applications and associated data analysis.DNA sequencing is an experimental process of determining the order of nucleotides in a given DNA molecule. Precise knowledge of the DNA sequence of entire genomes is paramount in identifying downstream genes, transcripts, and proteins, and in the consequent elucidation of biochemical processes taking place in a given organism. Until a decade ago, first-generation sequencing was dominated by the chain termination method developed by Fred Sanger, which generated detailed knowledge about the nucleotide sequence of many prokaryotic and several eukaryotic genomes, contributing to various genotyping and mutation discovery projects as well as assessing gene expression levels through serial analysis of gene expression. The most spectacular achievement, however, was the sequencing of the entire human genome, which took a decade and came at a cumulative cost of $2.7 billion before it was finally completed in 2003.