Exploration of methods for many-site genome editing with MAGE

Abstract

In order to modify bacterial genetic codes, modifications must be made throughout the genome wherever the affected codon is used. Making such numerous and widespread genomic changes requires specialized techniques. MAGE is well-suited to this task, being highly amenable to multiplexing and having low time and resource costs per site. MAGE has been used as a first stage in recoding efforts, converting small clusters of sites in separate strains to be combined by other means, but improvements in MAGE technique suggest the possibility of using it to produce fully-recoded strains directly. To this end, I compare strategies based on co-selected MAGE and apply the best by performing 80 site conversions spread over 1/4 of the E coli genome.