A Multiplex Genome Editing Method for Escherichia coli Based on CRISPR-Cas12a.

ABSTRACT: Various methods for editing specific sites in the Escherichia coli chromosome are available, and gene-size (∼1 kb) integration into a single site or to introduce deletions, short insertions or point mutations into multiple sites can be conducted in a short period of time. However, a method for rapidly integrating multiple gene-size sequences into different sites has not been developed yet. Here, we describe a method and plasmid system that makes it possible to simultaneously insert genes into multiple specific loci of the E. coli genome without the need for chromosomal markers. The method uses a CRISPR-Cas12a system to eliminate unmodified cells by double-stranded DNA cleavage in conjunction with the phage-derived λ-Red recombinases to facilitate recombination between the chr

SUBMITTER: Ao X 

PROVIDER: S-EPMC6189296 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature