清脆的
引导RNA
Cas9
核糖核酸
化学
基因组编辑
连接器
RNA编辑
亚基因组mRNA
基因
计算生物学
生物化学
生物
计算机科学
操作系统
作者
Dongyang Zhang,Luping Liu,Shuaijiang Jin,Ember M. Tota,Zijie Li,Xijun Piao,Xuan Zhang,Xiang‐Dong Fu,Neal K. Devaraj
摘要
Chemical cross-linking enables rapid identification of RNA-protein and RNA-nucleic acid inter- and intramolecular interactions. However, no method exists to site-specifically and covalently cross-link two user-defined sites within an RNA. Here, we develop RNA-CLAMP, which enables site-specific and enzymatic cross-linking (clamping) of two selected guanine residues within an RNA. Intramolecular clamping can disrupt normal RNA function, whereas subsequent photocleavage of the cross-linker restores activity. We used RNA-CLAMP to clamp two stem loops within the single-guide RNA (sgRNA) of the CRISPR-Cas9 gene editing system via a photocleavable cross-linker, completely inhibiting gene editing. Visible light irradiation cleaved the cross-linker and restored gene editing with high spatiotemporal resolution. Design of two photocleavable linkers responsive to different wavelengths of light allowed multiplexed photoactivation of gene editing in mammalian cells. This photoactivated CRISPR-Cas9 gene editing platform benefits from undetectable background activity, provides a choice of activation wavelengths, and has multiplexing capabilities.
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