Executing cell-specific cross-linking immunoprecipitation and sequencing (seCLIP) in C. elegans

Stephen M Blazie; Yishi Jin

doi:10.1016/j.xpro.2022.101959

Executing cell-specific cross-linking immunoprecipitation and sequencing (seCLIP) in C. elegans

STAR Protoc. 2023 Mar 17;4(1):101959. doi: 10.1016/j.xpro.2022.101959. Epub 2022 Dec 24.

Authors

Stephen M Blazie¹, Yishi Jin²

Affiliations

¹ Department of Neurobiology, School of Biological Sciences, University of California, San Diego, La Jolla, CA 92093, USA. Electronic address: sblazie@ucsd.edu.
² Department of Neurobiology, School of Biological Sciences, University of California, San Diego, La Jolla, CA 92093, USA. Electronic address: yijin@ucsd.edu.

Abstract

The single-end enhanced cross-linking immunoprecipitation (seCLIP) method is well suited for efficient and unbiased transcriptome-wide interrogation of RNA-binding protein (RBP) interaction sites. Here, we provide a protocol for executing cell-specific seCLIP for any desired RBP in Caenorhabditis elegans. We begin with steps and recommendations for transgene construction and Cas9-mediated chromosomal integration. We provide detailed procedures for isolation of RBP-associated RNA fragments, subsequent library preparation, and sequencing. We further discuss best practices for data analysis, interpretation of results, and troubleshooting. For complete details on the use and execution of this protocol, please refer to Blazie et al. (2021).¹.

Keywords: Genetics; Genomics.

Publication types

Research Support, N.I.H., Extramural

MeSH terms

Animals
Binding Sites
Caenorhabditis elegans* / genetics
Caenorhabditis elegans* / metabolism
Immunoprecipitation
RNA-Binding Proteins* / genetics
RNA-Binding Proteins* / metabolism
Transcriptome

Substances

RNA-Binding Proteins

Abstract

Publication types

MeSH terms

Substances

Grants and funding