Targeted gene deletions in the dimorphic fungal pathogen Histoplasma using an optimized episomal CRISPR/Cas9 system.

ABSTRACT: The rapid development of CRISPR/CRISPR-associated (Cas) systems has revolutionized the ability to produce genetic mutations in a desired locus, particularly in organisms with low rates of homologous recombination. Histoplasma is an important respiratory and systemic fungal pathogen that has few reverse genetic options. We describe an optimized CRISPR/Cas system for the efficient generation of mutations in desired genes. The limited requirements for CRISPR/Cas, namely a gene-targeting guide RNA (gRNA) and expression of a Cas endonuclease, enabled both the gRNA and the Streptococcus pyogenes Cas9 gene to be expressed from a single episomal vector. The gRNAs are expressed from a strong Pol(II) promoter, a critical parameter for increasing the recovery of mutated genes, and proce

SUBMITTER: Rappleye CA 

PROVIDER: S-EPMC10449496 | biostudies-literature | 2023 Aug

REPOSITORIES: biostudies-literature