A simple proficient method for targeted genome editing in Escherichia coli

Abstract

The ability to precisely and seamlessly modify the targeted genome is needed for metabolic engineering and synthetic biology aiming to create potent bio-systems. Here, we report a promising method in Escherichia coli developed by combining short single stranded DNA (oligos)- or long double stranded DNA-mediated genome editing and TetA dual selection system. This method could be successfully and rapidly used for targeted genome engineering including deletions, insertions, replacement and point mutation without the inactivation of methyl-directed mismatch repair (MMR) system and plasmid cloning. It was observed that even single recombination event was sufficient to get the positive genome edited mutants with the efficiencies of over 90% (insertion) or 70% (replacement/deletion) by the optimized selection process on NiCl2.