用于在细胞系中下拉环状 RNA 结合蛋白的回拼接接头探针的设计和使用。

Design and use of a back splicing junction probe for pulldown of circular RNA-binding proteins in cell lines.

机构信息

Sunnybrook Research Institute, and Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON, Canada.

出版信息

STAR Protoc. 2022 Dec 16;3(4):101702. doi: 10.1016/j.xpro.2022.101702. Epub 2022 Sep 22.

DOI:10.1016/j.xpro.2022.101702

PMID:36149796

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC9519607/

Abstract

Due to the unique structure of circular RNAs, it is challenging to use traditional pulldown approaches. Here, we describe the design and use of a probe that spans the back splicing junction (BSJ), enabling interaction with circular RNAs. The probe repeats four times, allowing efficient and specific pulldown of circular RNAs and their binding partners. This protocol describes the steps for mouse cardiac fibroblast (MCF) cells; we have also verified the protocol in other cell types. For complete details on the use and execution of this protocol, please refer to Wu et al. (2021).

摘要

由于环状 RNA 的独特结构，使用传统的下拉方法具有挑战性。在这里，我们描述了一种探针的设计和使用方法，该探针跨越了后拼接连接点 (BSJ)，能够与环状 RNA 相互作用。该探针重复四次，能够有效地、特异性地下拉环状 RNA 及其结合伙伴。本方案描述了用于小鼠心肌成纤维细胞 (MCF) 的步骤；我们也已经在其他细胞类型中验证了该方案。如需了解该方案使用和执行的完整详细信息，请参考 Wu 等人 (2021)。