文献类型：期刊文献

英文题名：An Orthogonal CRISPR/dCas12a System for RNA Imaging in Live Cells

作者：Jia, Hai-Yan[1];Zhang, Xin-Yue[1];Ye, Bang-Ce[1,2];Yin, Bin-Cheng[1,2]

机构：[1]East China Univ Sci & Technol, Shanghai Collaborat Innovat Ctr Biomfg Technol, State Key Lab Bioreactor Engn, Lab Biosyst & Microanal, Shanghai 200237, Shanghai, Peoples R China;[2]Shihezi Univ, Sch Chem & Chem Engn, Shihezi 832000, Xinjiang, Peoples R China

年份：2024

卷号：96

期号：15

起止页码：5913

外文期刊名：ANALYTICAL CHEMISTRY

收录：;EI(收录号：20241415848341);WOS:【SCI-EXPANDED(收录号:WOS:001195924800001)】；

基金：This work was jointly supported by the National Natural Science Foundation of China (Grants 22374047, 22134003), and the Fundamental Research Funds for the Central Universities.

语种：英文

外文关键词：Oligonucleotides

摘要：CRISPR/Cas technology has made great progress in the field of live-cell imaging beyond genome editing. However, effective and easy-to-use CRISPR systems for labeling multiple RNAs of interest are still needed. Here, we engineered a CRISPR/dCas12a system that enables the specific recognition of the target RNA under the guidance of a PAM-presenting oligonucleotide (PAMmer) to mimic the PAM recognition mechanism for DNA substrates. We demonstrated the feasibility and specificity of this system for specifically visualizing endogenous mRNA. By leveraging dCas12a-mediated precursor CRISPR RNA (pre-crRNA) processing and the orthogonality of dCas12a from different bacteria, we further demonstrated the proposed system as a simple and versatile molecular toolkit for multiplexed imaging of different types of RNA transcripts in live cells with high specificity. This programmable dCas12a system not only broadens the RNA imaging toolbox but also facilitates diverse applications for RNA manipulation.