Streamlined high-throughput cloning protocol to generate arrayed mutant libraries.

Autor:	Sun KT; Department of Biochemistry, University of Alberta, Edmonton, AB, Canada. Electronic address: ksun4@ualberta.ca., Patel TS; Department of Biochemistry, University of Alberta, Edmonton, AB, Canada., Kim J; Department of Biochemistry, University of Alberta, Edmonton, AB, Canada., Tang HSH; Department of Biochemistry, University of Alberta, Edmonton, AB, Canada., Eskandari-Sedighi G; Department of Biochemistry, University of Alberta, Edmonton, AB, Canada., Sureshkumar H; Department of Biochemistry, University of Alberta, Edmonton, AB, Canada., Schieve D; Department of Biochemistry, University of Alberta, Edmonton, AB, Canada., Mok SA; Department of Biochemistry, University of Alberta, Edmonton, AB, Canada. Electronic address: sueann@ualberta.ca.
Jazyk:	angličtina
Zdroj:	STAR protocols [STAR Protoc] 2023 Mar 17; Vol. 4 (1), pp. 101930. Date of Electronic Publication: 2022 Dec 13.
DOI:	10.1016/j.xpro.2022.101930
Abstrakt:	Large-scale, site-directed mutagenesis enables rapid characterization of the biochemical and biological properties of proteins. Here, we present a cost-effective and adaptable cloning pipeline to generate arrayed gene libraries for a construct of interest. We detail steps to use an open access web app to automate the design of mutagenesis primers optimized for our cloning protocols in a 96-well plate format. The protocol allows most molecular biology labs to clone 96 mutants (from PCR to sequence ready plasmid) in 3 days. Competing Interests: Declaration of interests The authors declare no competing interests. (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)
Databáze:	MEDLINE
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