Adaptive Evolution of a Minimal Organism with a Synthetic Genome

The bacterial strain JCVI-syn3.0 stands as the first example of a living organism with a minimized synthetic genome, derived from the Mycoplasma mycoides genome and chemically synthesized in vitro. Here we report the experimental evolution of a syn3.0- derived strain. Ten independent replicates were evolved for several hundred generations, leading to growth rate improvements of >15%. Endpoint strains possessed an average of 8 mutations composed of indels and SNPs, with a pronounced C/G->A/T transversion bias. Multiple genes were repeated mutational targets across the independent lineages, including phase variable lipoprotein activation, 5 distinct nonsynonymous substitutions in the same membrane transporter protein, and inactivation of an uncharacterized gene. Transcriptomic analysis revealed an overall tradeoff reflected in upregulated ribosomal proteins and downregulated DNA and RNA related proteins during adaptation. This work establishes the suitability of synthetic, minimal strains for laboratory evolution, providing a means to optimize strain growth characteristics and elucidate gene functionality. Overall design: Comparitive expression analysis between evolved strains using RNA-seq