SHAPE to Probe RNA Structure and RNA-Protein Interactions In Vitro

Abstract

Selective 2' hydroxyl acylation analyzed by primer extension (SHAPE) is used to distinguish between the levels of flexibility of nucleotides regulated by base pairing or protein binding. In this method, a reagent reacts with the 2' hydroxyl group to form an adduct, which is then detected by reverse transcription reaction. The number of RNA molecules with an adduct at a specific nucleotide position indicates the SHAPE reactivity of that nucleotide. Here, we describe the method for probing the structure of an RNA in a protein-free or a protein-bound state by in vitro SHAPE.

Keywords: Phosphorimaging; RNA end-labeling; RNA structure; RNA–protein interaction; Reverse transcription; Selective 2′ hydroxyl acylation analyzed by primer extension (SHAPE); Sequencing gel.