Functional genomic screening in Komagataella phaffii enabled by high-activity CRISPR-Cas9 library.

CRISPR-based high-throughput genome-wide loss-of-function screens are a valuable approach to functional genetics and strain engineering. The yeast Komagataella phaffii is a host of particular interest in the biopharmaceutical industry and as a metabolic engineering host for proteins and metabolites. Here, we design and validate a highly active 6-fold coverage genome-wide sgRNA library for this biotechnologically important yeast containing 30,848 active sgRNAs targeting over 99% of its coding sequences.

acCRISPR: an activity-correction method for improving the accuracy of CRISPR screens.

High throughput CRISPR screens are revolutionizing the way scientists unravel the genetic underpinnings of engineered and evolved phenotypes. One of the critical challenges in accurately assessing screening outcomes is accounting for the variability in sgRNA cutting efficiency. Poorly active guides targeting genes essential to screening conditions obscure the growth defects that are expected from disrupting them.