Chapter 6 - RNA-Sequencing and Methylome Analysis

Abstract

In addition to measuring gene expression levels, next-generation sequencing (NGS)-based RNA-sequencing methods (RNA-seq) can also reveal unknown transcripts and splicing isoforms, and provide quantitative measurements of alternatively spliced isoforms. The total RNA complement of a cell or a population of cells, also known as transcriptome, can be analyzed by this technology. RNA-seq greatly extends the possibilities of transcriptome studies to the analysis of gene isoforms, translocation events, nucleotide variations, and posttranscriptional base modifications. Due to its flexibility and the declining cost of massively parallel sequencing, NGS-based RNA-seq is becoming an increasingly attractive tool to investigate the full range of transcripts and to reveal the complex landscape and dynamics of the transcriptome. However, significant challenges remain for RNA-seq to migrate into clinical practice for diagnostic and predictive applications. This chapter discusses the concepts and considerations of RNA-seq and methylome sequencing, and describes the basic approaches and challenges associated with these applications.