Domesticating the Genome of a Naturally Transformable Bacterium

As the field of synthetic biology moves towards more ambitious genome engineering efforts, the need for chassis organisms better suited for genome modification is becoming more apparent. The bacterium Acinetobacter baylyi ADP1 could provide an ideal next-generation synthetic biology chassis due to its natural transformability. We conducted a 1000-generation adaptive evolution experiment in ADP1 to study the stability of this strain. We observed the gradual loss of transformability over time and, furthermore, observed the emergence of a high-aggregation phenotype. The genomes of evolved clones were sequenced to determine the genetic basis of these phenotypic changes. We found that
the activity of ADP1's sole IS-element family IS1236 was the main driver behind these changes and also observed the loss of a 49-kb prophage region that had been previously used as a landing site for integrating heterologous genes into the chromosome. We deleted all copies of IS1236 from the ADP1 genome to create an IS-less strain (ADP1-ISx) with increased genetic stability. This strain showed a much greater decrease in the frequency of gene-inactivating mutations than expected from the rates of spontaneous IS1236 insertions. Unexpectedly, ISx also has improved transformability and a reduced rate of autolysis. In sum, our work demonstrates the utility of 'domesticating' the genomes of microbial strains by removing selfish DNA elements and has generated a new bacterial chassis (ADP1-ISx) with improved transformability and genomic stability for future genome engineering efforts.