Transcriptome-Wide Mapping of N⁶-Methyladenosine by m⁶A-Seq

Methods Enzymol. 2015:560:131-47. doi: 10.1016/bs.mie.2015.03.001. Epub 2015 Jun 2.

Abstract

A detailed protocol for isolation and sequencing of an enriched population of m(6)A-methylated RNA fragments to create m(6)A methylome maps is outlined. Our approach was developed to fill a void that existed because of a lack of methods for the detection of m(6)A in RNA in an unbiased, high-throughput, and high-resolution manner. This method integrates immunoprecipitation of methylated, randomly fragmented RNA using a highly specific anti-m(6)A antibody to obtain an enriched population of modified fragments and massively parallel sequencing, resulting in mapping of this modification throughout the transcriptome.

Keywords: Immunocapturing; N(6)-Methyladenosine; Next-generation sequencing; RNA immunoprecipitation; RNA methylation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine / analogs & derivatives*
  • Adenosine / chemistry
  • Adenosine / genetics
  • Adenosine / isolation & purification
  • High-Throughput Nucleotide Sequencing / methods*
  • Methylation
  • RNA / chemistry*
  • RNA / genetics
  • Transcriptome / genetics*

Substances

  • RNA
  • N-methyladenosine
  • Adenosine