A detailed protocol for expression, purification, and activity determination of recombinant SaCas9

Recombinant SaCas9 is useful for a broad range of applications in the context of genome editing, especially when the specific protospacer adjacent motifs of other Cas9 derivatives are missing. Here, we describe a detailed protocol for the expression and purification of recombinant SaCas9. We detail the main steps for immobilized metal affinity chromatography and size exclusion chromatography. In addition, we present an assay for activity determination of SaCas9. Active SaCas9 can be purified in a week by using this protocol.

Metadaten
Author:	Franziska Flottmann GND, Greta Marie Pohl GND, Jan Gummert ORCiD GND, Hendrik Milting ORCiD GND, Andreas Brodehl ORCiD GND
URN:	urn:nbn:de:hbz:294-101726
DOI:	https://doi.org/10.1016/j.xpro.2022.101276
Parent Title (English):	STAR Protocols
Publisher:	Cell Press
Place of publication:	Cambridge
Document Type:	Article
Language:	English
Date of Publication (online):	2023/09/19
Date of first Publication:	2022/06/17
Publishing Institution:	Ruhr-Universität Bochum, Universitätsbibliothek
Tag:	Open Access Fonds
Volume:	3
Issue:	2, Article 101276
First Page:	101276-1
Last Page:	101276-14
Note:	Article Processing Charge funded by the Deutsche Forschungsgemeinschaft (DFG) and the Open Access Publication Fund of Ruhr-Universität Bochum.
Institutes/Facilities:	Herz- und Diabeteszentrum NRW
	Herz- und Diabeteszentrum NRW, Klinik für Thorax- und Kardiovaskularchirurgie
	Erich und Hanna Klessmann-Institut für kardiovaskuläre Forschung und Entwicklung
Dewey Decimal Classification:	Technik, Medizin, angewandte Wissenschaften / Medizin, Gesundheit
open_access (DINI-Set):	open_access
Licence (English):	Creative Commons - CC BY 4.0 - Attribution 4.0 International

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