Comprehensive Genome-wide Perturbations via CRISPR Adaptation Reveal Complex Genetics of Antibiotic Sensitivity.

ABSTRACT: Genome-wide CRISPR screens enable systematic interrogation of gene function. However, guide RNA libraries are costly to synthesize, and their limited diversity compromises the sensitivity of CRISPR screens. Using the Streptococcus pyogenes CRISPR-Cas adaptation machinery, we developed CRISPR adaptation-mediated library manufacturing (CALM), which turns bacterial cells into "factories" for generating hundreds of thousands of crRNAs covering 95% of all targetable genomic sites. With an average gene targeted by more than 100 distinct crRNAs, these highly comprehensive CRISPRi libraries produced varying degrees of transcriptional repression critical for uncovering novel antibiotic resistance determinants. Furthermore, by iterating CRISPR adaptation, we rapidly generated dual-crRNA libraries re

SUBMITTER: Jiang W 

PROVIDER: S-EPMC7169367 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature