Abstract

Selective 2'-hydroxyl acylation analyzed by primer extension (SHAPE) structure probing techniques characterize the secondary structure of RNA molecules, which influence their functions and interactions. A variation of SHAPE, footprinting SHAPE (fSHAPE), probes RNA in the presence and absence of protein to identify RNA bases that hydrogen-bond with protein. SHAPE or fSHAPE coupled with enhanced crosslinking and immunoprecipitation (SHAPE-eCLIP or fSHAPE-eCLIP) pulls down RNAs bound by any protein of interest and returns their structure or protein interaction information, respectively. Here, we describe detailed protocols for SHAPE-eCLIP and fSHAPE-eCLIP and an analysis protocol for fSHAPE. For complete details on the use and execution of these protocols, please refer to Corley et al. (2020).