CRISPR Guide RNA Design: Methods and Protocols

by Ferry, Quentin R. V. Fulga, Tudor A. Knapp, David J. H. F.

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Title Statements

Full title :CRISPR Guide RNA Design : Methods and Protocols / Edited by Tudor A. Fulga, David J.H.F. Knapp, and Quentin R.V. Ferry (Radcliffe Department of Medicine, Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK).

Summary

This detailed volume focuses on the CRISPR-associated guide RNA and how it can be designed, modified, and validated for a broad repertoire of purposes. Beginning with a section on computational design of target-specific guide RNAs, the book continues by covering chemical modifications to alter guide RNA stability, specificity, and efficiency, as well as to create inducible guide RNAs, append additional functional domains, and express guide RNAs in a conditional manner. It concludes with methods for measuring off-target guide RNA activity. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and essential, CRISPR Guide RNA Design: Methods and Protocols provides a comprehensive pipeline for guide RNA design and aims to be an invaluable resource in applying this powerful technology to basic research and therapeutic applications.

Edition Statements

Publication Date:2021

Publication Statements

Publisher(s):Humana Press
Place of Publication:New York, NY
Language:eng
Publication Date:2021

Notes

Part I: In Silico Design and Optimization of Guide RNA Sequences; ; 1. Cloud-Based Design of Short Guide RNA (sgRNA) Libraries for CRISPR Experiments; Florian Heigwer and Michael Boutros; ; 2. Web-Based CRISPR Toolkits: Cas-OFFinder, Cas-Designer, and Cas-Analyzer; Gue-Ho Hwang, Jin-Soo Kim, and Sangsu Bae; ; Part II: Chemically-Modified Guide RNAs; ; 3. Chemical Modification of Guide RNAs for Improved CRISPR Activity in CD34+ Human Hematopoietic Stem and Progenitor Cells; Jenny Shapiro, Adi Tovin, Ortal Iancu, Daniel Allen, and Ayal Hendel; ; 4. Gene Disruption Using Chemically-Modified CRISPR-Cpf1 RNA; Moira A. McMahon and Meghdad Rahdar; ; 5. 'Split-and-Click' sgRNA; Lapatrada Taemaitree, Arun Shivalingam, Afaf H. El-Sagheer, and Tom Brown; ; 6. Chimeric DNA-RNA Guide RNA Designs; Shuhan Lu, Ying Zhang, and Hao Yin; ; Part III: Expanding the CRISPR Toolbox ; ; 7. .

- Harnessing tRNA for Processing Ability and Promoter Activity; David J.H.F. Knapp and Tudor A. Fulga; ; 8. Targeting Noncoding RNA Domains to Genomic Loci with CRISPR-Display: Guidelines for Designing, Building, and Testing sgRNA-ncRNA Expression Constructs; David M. Shechner; ; 9. Controlling the Activity of CRISPR Transcriptional Regulators with Inducible sgRNAs; Quentin R.V. Ferry; ; 10. Gene Manipulation Using Fusion Guide RNAs for Cas9 and Cas12a; Ha Rim Shin, Jiyeon Kweon, and Yongsub Kim; ; Part IV: Characterization of CRISPR Efficacy and Specificity; ; 11. Methods for Measuring CRISPR/Cas9 DNA Cleavage in Cells; Christopher R. Cromwell, Juan Jovel, and Basil P. Hubbard; ; 12. In Vitro Assays for Comparing the Specificity of First- and Next-Generation CRISPR/Cas9 Systems; Christopher R. Cromwell and Basil P. Hubbard; ; 13. .

- Profiling Genome-Wide Specificity of CRISPR-Cas9 Using Digenome-Seq; Daesik Kim and Jin-Soo Kim; ; 14. Detection of CRISPR/Cas9-Generated Off-Target Effect by Integration-Defective Lentiviral Vector; Xiaoling Wang, Youjun Wu, and Jiing-Kuan Yee; ; 15. Genome-Wide CRISPR Off-Target DNA Break Detection by the BLISS Method; Roberto Ballarino, Britta A.M. Bouwman, and Nicola Crosetto.