Protocol for detecting lncRNA-protein interactions in vitro by tRSA RNA pull-down assay

Liyun Jiang; Jun Yang; Reqing He; Youlin Zhu; Dong Wang

doi:10.1016/j.xpro.2023.102818

Protocol for detecting lncRNA-protein interactions in vitro by tRSA RNA pull-down assay

STAR Protoc. 2024 Mar 15;5(1):102818. doi: 10.1016/j.xpro.2023.102818. Epub 2024 Jan 5.

Authors

Liyun Jiang¹, Jun Yang¹, Reqing He¹, Youlin Zhu¹, Dong Wang²

Affiliations

¹ Key Laboratory of Molecular Biology and Gene Engineering in Jiangxi Province, College of Life Science, Nanchang University, Jiangxi 330031, China.
² Key Laboratory of Molecular Biology and Gene Engineering in Jiangxi Province, College of Life Science, Nanchang University, Jiangxi 330031, China. Electronic address: dongwang@ncu.edu.cn.

Abstract

Long non-coding RNAs (lncRNAs) work together with diverse RNA-binding proteins (RBPs) to fulfill key regulations in important cellular functions. Here, we present a protocol to detect lncRNA-RBP interactions in vitro using a tRNA scaffold containing a streptavidin aptamer pull-down assay. We describe steps for preparing both protein and lncRNA transcripts, lncRNA-protein interaction detection with an in vitro binding assay, and western blot analysis. This protocol is applicable to screen for RNA-interacting proteins using cell lysates followed by mass spectrometry analysis. For complete details on the use and execution of this protocol, please refer to Yang et al. (2023).¹.

Keywords: Gene Expression; Molecular Biology; Plant sciences.

MeSH terms

RNA, Long Noncoding* / genetics
RNA, Transfer

Substances

RNA, Long Noncoding
RNA, Transfer