RNA Sequencing Protocols for Short-Read Sequencing

Laura Vasquez-Velez, Veera D’Mello, Patricia Soteropoulos

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

RNA sequencing (RNA-seq) methodologies allow the discovery of novel variants and transcripts. These comprise three general steps: (1) capture of RNA species of interest, (2) conversion of RNA to complementary DNA (cDNA), and (3) modification of cDNA to fit the sequencing platform. Here we describe four different library preparation protocols for short-read sequencing: cDNA synthesis with poly(A) selection, library preparation with ribosomal depletion, and cDNA synthesis with SMART® (Switching Mechanism at 5′ end of RNA Template) technology for low and Pico inputs.

Original languageEnglish (US)
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Pages125-158
Number of pages34
DOIs
StatePublished - 2025

Publication series

NameMethods in Molecular Biology
Volume2866
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Genetics

Keywords

  • Library preparation
  • Poly(A) selection
  • RNA-seq
  • Ribosomal depletion
  • Short-read sequencing
  • cDNA

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