RNA structure probing to characterize RNA-protein interactions on a low abundance pre-mRNA in living cells

RNA. 2020 Dec 11;27(3):343-358. doi: 10.1261/rna.077263.120. Online ahead of print.


In vivo RNA structure analysis has become a powerful tool in molecular biology, largely due to the coupling of an increasingly diverse set of chemical approaches with high-throughput sequencing. This has resulted in a transition from single target to transcriptome-wide approaches. However, these methods require sequencing depths that preclude studying low abundance targets, which are not sufficiently captured in transcriptome-wide approaches. Here we present a ligation-free method to enrich for low abundance RNA sequences, which improves the diversity of molecules analyzed and results in improved analysis. In addition, this method is compatible with any choice of chemical adduct or read-out approach. We utilized this approach to study an autoregulated event in the pre-mRNA of the splicing factor, muscleblind-like splicing regulator 1 (MBNL1).

Keywords: RNA structure; SHAPE-MaP; muscleblind; ultraconserved elements.