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RiboMeth-seq: Profiling of 2′-O-Me in RNA

  • Nicolai Krogh
  • Ulf Birkedal
  • Henrik NielsenEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1562)

Abstract

RiboMeth-seq is a sequencing-based method for mapping and quantitation of one of the most abundant RNA modifications, ribose methylation. It is based on a simple chemical principle, namely the several orders of magnitude difference in nucleophilicity of a 2′-OH and a 2-O-Me. Thus, the method combines alkaline fragmentation and a specialized library construction protocol based on 5-OH and 2,3 cyclic phosphate ends to prepare RNA for sequencing. The read-ends of library fragments are used for mapping with nucleotide resolution and calculation of the fraction of molecules methylated at the 2-O-Me sites.

Key words

2-O-Me Ribose methylation Box C/D snoRNA Ribosome Ribosomal RNA RNA library RNA-seq 

Notes

Acknowledgments

The work was supported by the Danish Research Council for Independent Research and the Danish Cancer Research Foundation.

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Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  1. 1.Department of Cellular and Molecular MedicineThe Panum Institute, University of CopenhagenCopenhagenDenmark

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