CRISPR-Cas9 ribonucleoprotein-mediated co-editing and counterselection in the rice blast fungus.

ABSTRACT: The rice blast fungus Magnaporthe oryzae is the most serious pathogen of cultivated rice and a significant threat to global food security. To accelerate targeted mutation and specific genome editing in this species, we have developed a rapid plasmid-free CRISPR-Cas9-based genome editing method. We show that stable expression of Cas9 is highly toxic to M. oryzae. However efficient gene editing can be achieved by transient introduction of purified Cas9 pre-complexed to RNA guides to form ribonucleoproteins (RNPs). When used in combination with oligonucleotide or PCR-generated donor DNAs, generation of strains with specific base pair edits, in-locus gene replacements, or multiple gene edits, is very rapid and straightforward. We demonstrate a co-editing strategy for the creation of single nuc

SUBMITTER: Foster AJ 

PROVIDER: S-EPMC6156577 | biostudies-literature | 2018 Sep

REPOSITORIES: biostudies-literature