Epitranscriptomic mapping of RNA modifications at single-nucleotide resolution using rhodamine sequencing (Rho-seq)

STAR Protoc. 2022 May 5;3(2):101369. doi: 10.1016/j.xpro.2022.101369. eCollection 2022 Jun 17.

Abstract

The recent development of epitranscriptomics revealed a new fundamental layer of gene expression, but the mapping of most RNA modifications remains technically challenging. Here, we describe our protocol for Rho-Seq, which enables the mapping of dihydrouridine RNA modification at single-nucleotide resolution. Rho-Seq relies on specific rhodamine-labeling of a subset of modified nucleotides that hinders reverse transcription. Although Rho-Seq was initially applied to the detection of dihydrouridine, we show here that it is applicable to other modifications including 7-methylguanosine or 4-thiouridine. For complete details on the use and execution of this protocol, please refer to Finet et al. (2022).

Keywords: Gene Expression; Molecular Biology; RNAseq; Sequence analysis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Nucleotides*
  • RNA / genetics
  • Rhodamines
  • Sequence Analysis, RNA / methods
  • Thiouridine*

Substances

  • Nucleotides
  • Rhodamines
  • Thiouridine
  • RNA