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Transcriptome-Wide Detection of 5-Methylcytosine by Bisulfite Sequencing

  • Thomas Amort
  • Xueguang Sun
  • Daria Khokhlova-Cubberley
  • Alexandra LusserEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1562)

Abstract

While low-throughput RNA bisulfite sequencing is the method of choice to assess the methylation status of specific cytosines in candidate RNAs, the combination of bisulfite treatment of RNA with today’s high-throughput sequencing techniques opens the door to methylation studies at nucleotide resolution on a transcriptome-wide scale. Below we describe a protocol for the transcriptome-wide analysis of total or fractionated poly(A)RNA in cells and tissues. Although the nature of the bisulfite sequencing protocol makes it comparably easy to translate from a low to a high-throughput approach, several critical points require attention before starting such a project. We describe a step-by-step protocol for planning and performing the experiment and analyzing the data.

Key words

Bisulfite sequencing RNA methylation 5-Methylcytosine High throughput Transcriptome Next-generation sequencing 

Notes

Acknowledgment

Funding in A.L’s lab is provided by the Austrian Science Fund (FWF): P27024-BBL.

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Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  • Thomas Amort
    • 1
  • Xueguang Sun
    • 2
  • Daria Khokhlova-Cubberley
    • 2
  • Alexandra Lusser
    • 1
    Email author
  1. 1.Division of Molecular Biology, BiocenterMedical University of InnsbruckInnsbruckAustria
  2. 2.Zymo Research CorpIrvineUSA

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