CRISPR, Softcover reprint of the original 1st ed. 2015
Methods and Protocols
Methods in Molecular Biology Series, Vol. 1311

This volume presents a list of cutting-edge protocols for the study of CRISPR-Cas defense systems and their applications at the genomic, genetic, biochemical and structural levels. CRISPR: Methods and Protocols guides readers through techniques that have been developed specifically for the analysis of CRISPR-Cas and techniques adapted from standard protocols of DNA, RNA and protein biology. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.

Authoritative and cutting-edge, CRISPR: Methods and Protocols provides a broad list of tools and techniques to study the interdisciplinary aspects of the prokaryotic CRISPR-Cas defense systems.

1. Investigating CRISPR RNA Biogenesis and Function using RNA-seq

Nadja Heidrich, Gaurav Dugar, Jörg Vogel, and Cynthia M. Sharma

2. In vitro Co-reconstitution of Cas Protein Complexes

André Plagens and Lennart Randau

3. Analysis of CRISPR pre-crRNA Cleavage

Erin L. Garside and Andrew M. MacMillan

4. Annotation and Classification of CRISPR-Cas Systems

Kira S. Makarova and Eugene V. Koonin

5. Computational Detection of CRISPR/crRNA Targets

Ambarish Biswas, Peter C. Fineran, and Chris M. Brown

6. High-throughput CRISPR typing of Mycobacterium tuberculosis Complex and Salmonella enterica Serotype Typhimurium

Christophe Sola, Edgar Abadia, Simon Le Hello, and François-Xavier Weill

7. Spacer-based Macroarrays for CRISPR Genotyping

Igor Mokrousov and Nalin Rastogi

8. Analysis of crRNA using Liquid Chromatography Electrospray Ionisation Mass Spectrometry (LC ESI MS)

Sakharam P. Waghmare, Alison O. Nwokeoji, and Mark J. Dickman

9. Rapid Multiplex Creation of Escherichia coli Strains Capable of Interfering with Phage Infection through CRISPR

Alexandra Strotskaia, Ekaterina Semenova, Ekaterina Savitskaya, and Konstantin Severinov

10. Exploring CRISPR Interference by Transformation with Plasmid Mixtures: Identification Of Target Interference Motifs in Escherichia coli

Cristóbal Almendros and Francisco J.M. Mojica

11. Electrophoretic Mobility Shift Assay of DNA and CRISPR-Cas Ribonucleoprotein Complexes

Tim Künne, Edze R. Westra^, and Stan J.J. Brouns

12. Expression and Purification of the CMR (Type III-B) Complex in Sulfolobus solfataricus

Jing Zhang and Malcolm F White

13. Procedures for Generating CRISPR Mutants With Novel Spacers Acquired from Viruses Or Plasmids

Marie-Ève Dupuis, Rodolphe Barrangou, and Sylvain Moineau

14. Archaeal Viruses of the Sulfolobales: Isolation, Infection, and CRISPR Spacer Acquisition

Susanne Erdmann and Roger A. Garrett

15. Using the CRISPR-Cas system to Positively Select Mutants In Genes Essential for its Function

Ido Yosef, Moran G. Goren, Rotem Edgar, and Udi Qimron

16. Analysis of Nuclease Activity of Cas1 Proteins Against Complex DNA Substrates

Natalia Beloglazova, Sofia Lemak, Robert Flick, and Alexander F. Yakunin

17. Characterizing Metal-Dependent Nucleases of CRISPR-Cas prokaryotic Adaptive Immunity Systems

Ki H. Nam, Matthew P. DeLisa^,  and Ailong Ke

18. Cas3 Nuclease-Helicase Activity Assays

Tomas Sinkunas, Giedrius Gasiunas,  and Virginijus Siksnys

19. Chemical and Enzymatic Footprint Analyses of R-loop Formation by Cascade-crRNA Complex

Ümit Pul

20. Creation and Analysis of a Virome: Using CRISPR Spacers

Michelle Davison and Devaki Bhaya

21. Targeted Mutagenesis in Zebrafish using CRISPR RNA-guided Nucleases

Woong Y Hwang, Yanfang Fu, Deepak Reyon, Andrew PW Gonzales, J Keith Joung, and Jing-Ruey Joanna Yeh

22. Precise Genome Editing of Drosophila with CRISPR RNA-guided Cas9

Scott J. Gratz, Melissa M. Harrison, Jill Wildonger, and Kate M. O’Connor-Giles

23. Targeted Transcriptional Repression in Bacteria using CRISPR Interference (CRISPRi)

John S. Hawkins, Spencer Wong, Jason M. Peters, Ricardo Almeida, and Lei S. Qi

Includes cutting-edge methods and protocols

Provides step-by-step detail essential for reproducible results

Contains key notes and implementation advice from the experts