Defining Transcribed Regions Using RNA-seq

Overview

Journal Nat Protoc

Specialties Biology
Pathology
Science

Date 2010 Feb 6

PMID 20134426

Citations 42

Authors

Brian T Wilhelm

Samuel Marguerat

Ian Goodhead

Jurg Bahler

Affiliations

Soon will be listed here.

Abstract

Next-generation sequencing technologies are revolutionizing genomics research. It is now possible to generate gigabase pairs of DNA sequence within a week without time-consuming cloning or massive infrastructure. This technology has recently been applied to the development of 'RNA-seq' techniques for sequencing cDNA from various organisms, with the goal of characterizing entire transcriptomes. These methods provide unprecedented resolution and depth of data, enabling simultaneous quantification of gene expression, discovery of novel transcripts and exons, and measurement of splicing efficiency. We present here a validated protocol for nonstrand-specific transcriptome sequencing via RNA-seq, describing the library preparation process and outlining the bioinformatic analysis procedure. While sample preparation and sequencing take a fairly short period of time (1-2 weeks), the downstream analysis is by far the most challenging and time-consuming aspect and can take weeks to months, depending on the experimental objectives.

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