Development of a CRISPR/Cas9-Based Tool for Gene Deletion in Issatchenkia orientalis
Microbial production of fuels and chemicals from renewable and readily available biomass is a sustainable and economically attractive alternative to petroleum-based production. Because of its unusual tolerance to highly acidic conditions, I. orientalis is a promising potential candidate for the manufacture of valued organic acids. Nevertheless, reliable and efficient genetic engineering tools in I. orientalis are limited. The results outlined in this paper describe a stable episomal ARS-containing plasmid and the first CRISPR/Cas9-based system for gene disruptions in I. orientalis , paving the way for applying genome engineering and metabolic engineering strategies and tools in this microorganism for production of fuels and chemicals.