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Genome-Wide Location Analyses of N6-Methyladenosine Modifications (m6A-Seq)

  • Benoit MolinieEmail author
  • Cosmas C. GiallourakisEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1562)

Abstract

N6-methyladenosine–sequencing (m6A-seq) is a critical tool to obtain an unbiased genome-wide picture of m6A sites of modification at high resolution. It allows the study of the impact of various perturbations on m6A modification distribution and the study of m6A functions. Herein, we describe the m6A-seq protocol, which entails RNA immunoprecipitation (RIP) performed on fragmented poly(A) RNA utilizing anti-m6A antibodies. The captured/enriched m6A positive RNA fragments are subsequently sequenced by RNA-seq in parallel with background control non-immunoprecipitated input RNA fragments. Analyses reveal peaks of m6A enrichment containing sites of modifications analogous to chromatin modification immunoprecipitation experiments.

Key words

m6A-Seq Epitranscriptome N6-Methyladenosine Genome-wide METTL3 METTL14 

Notes

Acknowledgments

This work was supported by MGH Start-up funds to Cosmas Giallourakis. We thank Yi Xing, Ph.D. and Jinkai Wang, Ph.D. at UCLA who have been our computational biologist collaborators on m6A related projects.

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Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  1. 1.Gastrointestinal UnitBostonUSA
  2. 2.Gastrointestinal Unit, Massachusetts General HospitalHarvard Medical SchoolBostonUSA

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