Mapping and Quantification of tRNA 2'-O-Methylation by RiboMethSeq.

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.