Improved vectors for retron-mediated CRISPR-Cas9 genome editing in Saccharomyces cerevisiae.

ABSTRACT: In vivo site-directed mutagenesis is a powerful genetic tool for testing the effects of specific alleles in their normal genomic context. While the budding yeast Saccharomyces cerevisiae possesses classical tools for site-directed mutagenesis, more efficient recent CRISPR-based approaches use Cas "cutting" combined with homologous recombination of a "repair" template that introduces the desired edit. However, current approaches are limited for fully prototrophic yeast strains and rely on relatively low-efficiency cloning of short gRNAs. We were thus motivated to simplify the process by combining the gRNA and its cognate repair template in cis on a single oligonucleotide. Moreover, we wished to take advantage of a new approach that uses an Escherichia coli retron (EcRT) to amplify repair te

SUBMITTER: Stuecker TN 

PROVIDER: S-EPMC12506651 | biostudies-literature | 2025 Oct

REPOSITORIES: biostudies-literature