CRISPR-based screening of genomic island excision events in bacteria

Significance The development of Clustered regularly interspaced short palindromic repeats (CRISPR)–CRISPR-associated genes (CAS)–based technology for targeted genome editing has revolutionized molecular biology approaches, but significant and outstanding gaps exist for applications in bacteria, the native hosts of these adaptive immune systems. This study shows that CRISPR-Cas systems can be directed to target and delete genomic islands that are flanked by insertion-sequence elements and devoid of essential genes. Naturally occurring minor subpopulations harboring deletions in genomic islands were identified and readily isolated using CRISPR-Cas screening. Promising applications of this approach can define minimal bacterial genomes, determine essential genes, and characterize genetically heterogeneous bacterial populations.