Harnessing CRISPR–Cas systems for bacterial genome editing

• CRISPR–Cas provides DNA-encoded, RNA-mediated sequence-specific targeting of DNA.
• The Cas9 endonuclease can be reprogrammed to cleave DNA precisely and generate sequence edits using the native DNA repair machinery.
• CRISPR–Cas-derived genetic tools afford a next-generation method for programmable and high-throughput functional genomics in bacteria.
• CRISPR–Cas technology may be exploited to target genomic features of interest in bacteria, opening new avenues for a wide array of genome engineering applications.

Manipulation of genomic sequences facilitates the identification and characterization of key genetic determinants in the investigation of biological processes. Genome editing via clustered regularly interspaced short palindromic repeats (CRISPR)–CRISPR-associated (Cas) constitutes a next-generation method for programmable and high-throughput functional genomics. CRISPR–Cas systems are readily reprogrammed to induce sequence-specific DNA breaks at target loci, resulting in fixed mutations via host-dependent DNA repair mechanisms. Although bacterial genome editing is a relatively unexplored and underrepresented application of CRISPR–Cas systems, recent studies provide valuable insights for the widespread future implementation of this technology. This review summarizes recent progress in bacterial genome editing and identifies fundamental genetic and phenotypic outcomes of CRISPR targeting in bacteria, in the context of tool development, genome homeostasis, and DNA repair.