通过 CRISPR-Cas13 系统对活细胞中的 RNA 进行动态成像的方案。
Protocol for Dynamic Imaging of RNA in Living Cells by CRISPR-Cas13 System.
机构信息
State Key Laboratory of Molecular Biology, Shanghai Key Laboratory of Molecular Andrology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, University of Chinese Academy of Sciences, Chinese Academy of Sciences, 320 Yueyang Road, Shanghai 200031, China.
School of Life Science and Technology, ShanghaiTech University, 100 Haike Road, Shanghai 201210, China.
出版信息
STAR Protoc. 2020 Jun 3;1(1):100037. doi: 10.1016/j.xpro.2020.100037. eCollection 2020 Jun 19.
This protocol uses endonuclease-dead, programmable RNA-guided RNA-targeting Cas13 RNases (d)Cas13 proteins fused with fluorescent proteins to visualize and track RNA dynamics in live cells. This protocol details several aspects of the procedure, including gRNA design, fluorescent protein selection, nuclear localization signal adjustment, raw data analysis, operation steps, and extended optional applications that have been successfully applied in the visualization of , , , and RNAs. For complete information on the use and execution of this protocol, please refer to Yang et al. (2019).
本方案使用与荧光蛋白融合的经酶切失活的、可编程的 RNA 导向 RNA 靶向 Cas13 RNA 酶（d）Cas13 蛋白，用于可视化和追踪活细胞内的 RNA 动态。本方案详细介绍了该程序的几个方面，包括 gRNA 设计、荧光蛋白选择、核定位信号调整、原始数据分析、操作步骤以及已成功应用于可视化 、 、 和 RNA 的扩展可选应用。如需了解该方案使用和执行的完整信息，请参考 Yang 等人（2019 年）的文献。
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