Protocol to process crosslinking and immunoprecipitation data into annotated binding sites

• Published in: STAR protocols 2024-06, Vol.5 (2), p.103040, Article 103040
• Main Authors: Xu, Shuhao, Nguyen, Grady G., Naritomi, Jack T., Kopalle, Hema M., Yee, Brian A., Rothamel, Katherine L., Boyle, Evan A., Yeo, Gene W.
• Format: Article
• Language: English
• Subjects: Binding Sites; Bioinformatics; Cross-Linking Reagents - chemistry; Gene Expression; Genomics; Humans; Immunoprecipitation - methods; RNA - genetics; RNA - metabolism; RNA-seq; Sequence analysis; Software
• ISSN: 2666-1667; 2666-1667
• DOI: 10.1016/j.xpro.2024.103040

Here, we present a protocol for using Skipper, a pipeline designed to process crosslinking and immunoprecipitation (CLIP) data into annotated binding sites. We describe steps for partitioning annotated transcript regions and fitting data to a beta-binomial model to call windows of enriched binding. From raw CLIP data, we detail how users can map reproducible RNA-binding sites to call enriched windows and perform downstream analysis. This protocol supports optional customizations for different use cases. For complete details on the use and execution of this protocol, please refer to Boyle et al.1 [Display omitted] •Protocol for applying a bioinformatics pipeline to process CLIP data•Conda-based and containerized software environments for simplifying pipeline setup•Steps for preparing pipeline resources using provided code snippets•Identification of increased true-positive RBP-RNA-binding sites in a few steps Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Here, we present a protocol for using Skipper, a pipeline designed to process crosslinking and immunoprecipitation (CLIP) data into annotated binding sites. We describe steps for partitioning annotated transcript regions and fitting data to a beta-binomial model to call windows of enriched binding. From raw CLIP data, we detail how users can map reproducible RNA-binding sites to call enriched windows and perform downstream analysis. This protocol supports optional customizations for different use cases.