Validating genome-wide CRISPR-Cas9 function in the non-conventional yeast Yarrowia lipolytica

摘要: Genome-wide mutational screens are central to understanding the genetic underpinnings of evolved and engineered phenotypes. The widespread adoption CRISPR-Cas9 genome editing has enabled such in many organisms, but identifying functional sgRNAs still remains a challenge. To address this limitation, we developed methodology quantify cutting efficiency each sgRNA genome-scale library biotechnologically important yeast Yarrowia lipolytica . Screening presence absence native DNA repair high-throughput quantification function leading identification high that cover 94% genes. Library validation enhanced classification essential genes by inactive guides create false negatives mask effects successful disruptions. Quantification guide effectiveness also creates dataset from which determinants can be identified. Finally, application identified mutations led lipid accumulation eliminated pseudohyphal morphology.