A highly precise and portable genome engineering method allows comparison of mutational effects across bacterial species.

ABSTRACT: Currently available tools for multiplex bacterial genome engineering are optimized for a few laboratory model strains, demand extensive prior modification of the host strain, and lead to the accumulation of numerous off-target modifications. Building on prior development of multiplex automated genome engineering (MAGE), our work addresses these problems in a single framework. Using a dominant-negative mutant protein of the methyl-directed mismatch repair (MMR) system, we achieved a transient suppression of DNA repair in Escherichia coli, which is necessary for efficient oligonucleotide integration. By integrating all necessary components into a broad-host vector, we developed a new workflow we term pORTMAGE. It allows efficient modification of multiple loci, without any observable off-targ

SUBMITTER: Nyerges A 

PROVIDER: S-EPMC4780621 | biostudies-literature | 2016 Mar

REPOSITORIES: biostudies-literature