Streamlined high-throughput cloning protocol to generate arrayed mutant libraries

ABSTRACT: Summary 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. Graphical abstract Highlights • Web app for automated generation of arrayed mutagenesis primer libraries• Optimized high-throughput site-directed mutagenesis PCR protocol• Streamlined bacterial transformation in a 96-well plate format• Automated magnetic-bead-bas

SUBMITTER: Sun K 

PROVIDER: S-EPMC9758493 | biostudies-literature | 2022 Dec

REPOSITORIES: biostudies-literature