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Methods Mol Biol. 2019;1956:283-303. doi: 10.1007/978-1-4939-9151-8_13.

RNA Sequencing in B-Cell Lymphomas.

Author information

1
Lymphoid Malignancies Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA.
2
Baylor College of Medicine, Houston, TX, USA.
3
Office of Intramural Research, Center for Information Technology, National Institutes of Health, Bethesda, MD, USA.
4
Institute of Pathology, University Hospital Giessen, Justus-Liebig-University Giessen, Giessen, Germany. roland.schmitz@patho.med.uni-giessen.de.

Abstract

High-throughput mRNA sequencing (RNA-Seq) provides both qualitative and quantitative evaluation of the transcriptome. This method uses complementary DNA (cDNA) to generate several millions of short sequence reads that are aligned to a reference genome allowing the comprehensive characterization of the transcripts in a cell. RNA-Seq has a wide variety of applications which lead to a pervasive adoption of this method well beyond the genomics community and a deployment of this technique as a standard part of the toolkit applied in life sciences. This chapter describes a protocol to perform mRNA sequencing using the Illumina NextSeq or MiSeq platforms, presents sequencing data quality metrics, and outlines a bioinformatic pipeline for sequence alignment, digital gene expression, identification of gene fusions, detection of transcript isoforms, description and annotation of genetic variants, and de novo immunoglobulin gene assembly.

KEYWORDS:

B cell; B-cell lymphoma; Gene expression; High-throughput sequencing; Immunoglobulin genes; Mutation; RNA-Seq; Transcriptome; VDJ

PMID:
30779040
DOI:
10.1007/978-1-4939-9151-8_13
[Indexed for MEDLINE]

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