Recyclable CRISPR/Cas9 mediated gene disruption and deletions in Histoplasma.

ABSTRACT: Targeted gene disruption is challenging in the dimorphic fungal pathogen Histoplasma due to the low frequency of homologous recombination. Transformed DNA is either integrated ectopically into the genome or maintained extra chromosomally by de novo addition of telomeric sequences. Based on a system developed in Blastomyces, we adapted a CRISPR/Cas9 system to facilitate targeted gene disruption in Histoplasma with high efficiency. We express a codon-optimized version of Cas9 as well as guide RNAs from a single ectopic vector carrying a selectable marker. Once the desired mutation is verified, one can screen for isolates that have lost the Cas9 vector by simply removing the selective pressure. Multiple mutations can then be generated in the same strain by retransf

SUBMITTER: Joehnk B 

PROVIDER: S-EPMC10350005 | biostudies-literature | 2023 Jul

REPOSITORIES: biostudies-literature