Profiling of RNA structure at single-nucleotide resolution using nextPARS

Abstract

RNA molecules play important roles in almost every cellular process, and their functions are mediated by their sequence and structure. Determining the secondary structure of RNAs is central to understanding RNA function and evolution. RNA structure probing techniques coupled to high-throughput sequencing allow determining structural features of RNA molecules at transcriptome-wide scales. Our group recently developed a novel Illumina-based implementation of in vitro parallel probing of RNA structures called nextPARS. Here, we describe a protocol for the computation of the nextPARS scores and their use to obtain the structural profile (single- or double-stranded state) of an RNA sequence at single-nucleotide resolution.