title: High-throughput RNA sequencing in B-cell lymphomas.
abstract: High-throughput mRNA sequencing (RNA-seq) uses massively parallel sequencing to allow an unbiased analysis of both genome-wide transcription levels and mutation status of a tumor. In the RNA-seq method, complementary DNA (cDNA) is used to generate short sequence reads by immobilizing millions of amplified DNA fragments onto a solid surface and performing the sequence reaction. The resulting sequences are aligned to a reference genome or transcript database to create a comprehensive description of the analyzed transcriptome. This chapter describes a protocol to perform RNA-seq using the Illumina sequencing platform, presents sequencing data quality metrics and outlines a bioinformatic pipeline for sequence alignment, digital gene expression, and mutation discovery.
authors.affiliation: Bioinformatics and Molecular Analysis Section, Division of Computational Bioscience, Center for Information Technology, National Institutes of Health, Bethesda, MD, USA.
authors.forename: Wenming
authors.initials: W
authors.lastname: Xiao
authors.forename: Bao
authors.initials: B
authors.lastname: Tran
authors.forename: Louis M
authors.initials: LM
authors.lastname: Staudt
authors.forename: Roland
authors.initials: R
authors.lastname: Schmitz
mesh_terms: D000818:Animals; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D016393:Lymphoma, B-Cell / Q000235:genetics*; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide / Q000235:genetics; D011786:Quality Control; D051381:Rats; D017423:Sequence Analysis, RNA / Q000379:methods*