Efficient and iterative retron-mediated in vivo recombineering in Escherichia coli.

ABSTRACT: Recombineering is an important tool in gene editing, enabling fast, precise and highly specific in vivo modification of microbial genomes. Oligonucleotide-mediated recombineering via the in vivo production of single-stranded DNA can overcome the limitations of traditional recombineering methods that rely on the exogenous delivery of editing templates. By modifying a previously reported plasmid-based system for fully in vivo single-stranded DNA recombineering, we demonstrate iterative editing of independent loci by utilizing a temperature-sensitive origin of replication for easy curing of the editing plasmid from recombinant cells. Optimization of the promoters driving the expression of the system's functional components, combined with targeted counterselection against

SUBMITTER: Ellington AJ 

PROVIDER: S-EPMC9165427 | biostudies-literature | 2022

REPOSITORIES: biostudies-literature