Improved vectors for retron-mediated CRISPR-Cas9 genome editing in <i>Saccharomyces cerevisiae</i>.

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 E. coli retron (EcRT)

SUBMITTER: Stuecker TN 

PROVIDER: S-EPMC11326209 | biostudies-literature | 2024 Aug

REPOSITORIES: biostudies-literature