Challenges in Reverse Engineering of Industrial Fermentation Strains

At Amyris, we optimize S. cerevisiae strains for industrial fermentations to manufacture biofuels and chemicals. Applying a combination of random mutagenesis, high-throughput Automated Strain Engineering (ASE), enzyme engineering, high-throughput phenotyping, and a tiered screening approach, we identify strains with improved yield, productivity, and product recovery. Strain “winners” serve as parents for further rounds of strain improvement, and are dissected using ‘omics technologies, whole genome sequencing, and identification of causal mutant alleles. We will discuss the merits, challenges and lessons learned from three different approaches to strain improvement.  First, we will discuss the relative contributions of mutagenic vs. rational genetic engineering approaches and the benefits of combining the two. Second, we will illustrate the use of whole genome sequencing and causal mutation identification to accelerate strain improvement. Third, we will describe attempts to reverse engineer top strains by building “clean” strains containing beneficial causal alleles and lacking “collateral damage” of mutagenesis.