Protocol for Dynamic Imaging of RNA in Living Cells by CRISPR-Cas13 System

Protocol for Dynamic Imaging of RNA in Living Cells by CRISPR-Cas13 System

Summary: 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 se...

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Main Authors: Yang Wang, Liang-Zhong Yang, Ling-Ling Chen
Format: Article
Language:English
Published: Elsevier 2020-06-01
Series:STAR Protocols
Online Access:http://www.sciencedirect.com/science/article/pii/S2666166720300241
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spelling doaj-faddcdc760a940d7a83f1579246066c92020-11-25T04:08:54ZengElsevierSTAR Protocols2666-16672020-06-0111100037Protocol for Dynamic Imaging of RNA in Living Cells by CRISPR-Cas13 SystemYang Wang0Liang-Zhong Yang1Ling-Ling Chen2State 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; Corresponding authorState 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, ChinaState 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; Corresponding authorSummary: 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 NEAT1, SatIII, MUC4, and GCN4 RNAs.For complete information on the use and execution of this protocol, please refer to Yang et al. (2019).http://www.sciencedirect.com/science/article/pii/S2666166720300241
collection DOAJ
language English
format Article
sources DOAJ
author Yang Wang
Liang-Zhong Yang
Ling-Ling Chen
spellingShingle Yang Wang
Liang-Zhong Yang
Ling-Ling Chen
Protocol for Dynamic Imaging of RNA in Living Cells by CRISPR-Cas13 System
STAR Protocols
author_facet Yang Wang
Liang-Zhong Yang
Ling-Ling Chen
author_sort Yang Wang
title Protocol for Dynamic Imaging of RNA in Living Cells by CRISPR-Cas13 System
title_short Protocol for Dynamic Imaging of RNA in Living Cells by CRISPR-Cas13 System
title_full Protocol for Dynamic Imaging of RNA in Living Cells by CRISPR-Cas13 System
title_fullStr Protocol for Dynamic Imaging of RNA in Living Cells by CRISPR-Cas13 System
title_full_unstemmed Protocol for Dynamic Imaging of RNA in Living Cells by CRISPR-Cas13 System
title_sort protocol for dynamic imaging of rna in living cells by crispr-cas13 system
publisher Elsevier
series STAR Protocols
issn 2666-1667
publishDate 2020-06-01
description Summary: 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 NEAT1, SatIII, MUC4, and GCN4 RNAs.For complete information on the use and execution of this protocol, please refer to Yang et al. (2019).
url http://www.sciencedirect.com/science/article/pii/S2666166720300241
work_keys_str_mv AT yangwang protocolfordynamicimagingofrnainlivingcellsbycrisprcas13system
AT liangzhongyang protocolfordynamicimagingofrnainlivingcellsbycrisprcas13system
AT linglingchen protocolfordynamicimagingofrnainlivingcellsbycrisprcas13system
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