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Mapping and Quantification of tRNA 2′-O-Methylation by RiboMethSeq

  • Adeline Galvanin
  • Lilia Ayadi
  • Mark Helm
  • Yuri Motorin
  • Virginie Marchand
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1870)

Abstract

Current development of epitranscriptomics field requires efficient experimental protocols for precise mapping and quantification of various modified nucleotides in RNA. Despite important advances in the field during the last 10 years, this task is still extremely laborious and time-consuming, even when high-throughput analytical approaches are employed. Moreover, only a very limited subset of RNA modifications can be detected and only rarely be quantified by these powerful techniques. In the past, we developed and successfully applied alkaline fragmentation-based RiboMethSeq approach for mapping and precise quantification of multiple 2′-O-methylation residues in ribosomal RNA. Here we describe a RiboMethSeq protocol adapted for the analysis of bacterial and eukaryotic tRNA species, which also contain 2′-O-methylations at functionally important RNA regions.

Key words

2′-O-Methylation High-throughput sequencing tRNA modification Ribose methylation Alkaline fragmentation 

Notes

Acknowledgement

This work was supported by joint ANR-DFG grant HTRNAMod (ANR-13-ISV8-0001/HE 3397/8-1) to MH and YM, and AO Lorraine University-Lorraine Region “Aberrant RNA methylation in cancer” funding to YM.

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Adeline Galvanin
    • 1
  • Lilia Ayadi
    • 1
  • Mark Helm
    • 2
  • Yuri Motorin
    • 1
    • 3
  • Virginie Marchand
    • 4
  1. 1.IMoPA UMR7365 CNRS-UL, BioPole Lorraine UniversityNancyFrance
  2. 2.Institute of Pharmacy and BiochemistryJohannes Gutenberg University MainzMainzGermany
  3. 3.Next-Generation Sequencing Core Facility, FR3209 BMCT, CNRS-UL, BioPole Lorraine UniversityNancyFrance
  4. 4.Next-Generation Sequencing Core Facility, UMS2008 IBSlor, CNRS-UL-INSERM, BioPole Lorraine UniversityNancyFrance

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