Mapping RNA Modifications Using Photo-Crosslinking-Assisted Modification Sequencing.

Abstract

Epitranscriptomic RNA modifications function as an important layer of gene regulation that modulates the function of RNA transcripts. A key step in understanding how RNA modifications regulate biological processes is the mapping of their locations, which is most commonly done by RNA immunoprecipitation (RIP) using modification-specific antibodies. Here, we describe the use of a photoactivatable ribonucleoside-enhanced cross-linking and immunoprecipitation (PAR-CLIP) method, in conjunction with RNA modification-specific antibodies, to map modification sites. First described as photo-crosslinking-assisted m6A sequencing (PA-m6A-seq), this method allows the mapping of RNA modifications at a higher resolution, with lower background than traditional RIP, and can be adapted to any RNA modification for which a specific antibody is available.