Deconvolution of in vivo protein-RNA contacts using fractionated eCLIP-seq

Abstract

Summary: Thousands of RNA-binding proteins orchestrate RNA processing and altered protein-RNA interactions frequently lead to disease. Here, we present experimental and computational analysis pipelines of fractionated eCLIP-seq (freCLIP-seq), a modification of enhanced UV-crosslinking and RNA immunoprecipitation followed by sequencing. FreCLIP-seq allows transcriptome-wide analysis of protein-RNA interactions at single-nucleotide level and provides an additional level of resolution by isolating binding signals of individual RNA-binding proteins within a multicomponent complex. Binding occupancy can be inferred from read counts and crosslinking events.For complete details on the use and execution of this protocol, please refer to Biancon et al. (2022). : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Keywords

• Bioinformatics
• Cell Biology
• Molecular Biology
• Protein Biochemistry
• Protein expression and purification
• RNAseq