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

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RNA Methylation

Part of the book series: Methods in Molecular Biology ((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.

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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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Authors and Affiliations

  1. Gastrointestinal Unit, 55 Fruit Street, Jackson 828, Boston, MA, 02114, USA

    Benoit Molinie

  2. Gastrointestinal Unit, Massachusetts General Hospital, Harvard Medical School, 55 Fruit Street, Jackson 828, Boston, MA, 02114, USA

    Cosmas C. Giallourakis

Authors
  1. Benoit Molinie
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  2. Cosmas C. Giallourakis
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Corresponding authors

Correspondence to Benoit Molinie or Cosmas C. Giallourakis .

Editor information

Editors and Affiliations

  1. Division of Molecular Biology, Biocenter, Medical University of Innsbruck, Innsbruck, Austria

    Alexandra Lusser

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Molinie, B., Giallourakis, C.C. (2017). Genome-Wide Location Analyses of N6-Methyladenosine Modifications (m6A-Seq). In: Lusser, A. (eds) RNA Methylation. Methods in Molecular Biology, vol 1562. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6807-7_4

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  • DOI: https://doi.org/10.1007/978-1-4939-6807-7_4

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-6805-3

  • Online ISBN: 978-1-4939-6807-7

  • eBook Packages: Springer Protocols

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