Genome minimized strains are obtained by removing genome segments associated with genes or processes either detrimental or unneeded under bioproduction conditions. They offer advantages as production chassis [1] by reducing transcriptional cost, eliminating competing functions and limiting unwanted regulatory interactions. Existing approaches for identifying stretches of DNA to remove are largely ad hoc based on information on presumably dispensable regions through experimentally determined non-essential genes and comparative genomics. As more sophisticated genome editing tools (e.g., CRISPR) are becoming commonplace, the need for a computational aid that will help successively minimize genomes consistent with a set of performance criteria beyond simply growth rate is becoming more pressing. Here we introduce a versatile genome reduction algorithm MinGenome that implements a mixed integer linear program (MILP) to iteratively identify the largest dispensable contiguous sequences without affecting the organism’s growth or other desirable traits. Known essential genes or genes that cause significant fitness or performance loss are flagged and their deletion is thus prohibited. MinGenome also preserves needed transcription factors and promoter regions ensuring that retained genes will be properly transcribed while also avoiding the simultaneous deletion of synthetic lethal pairs. The potential benefit of removing even larger contiguous stretches of DNA if only one or two essential genes (to be re-inserted elsewhere) are within the deleted sequence is explored. We apply the algorithm to design a minimized E. coli strain and a minimized B. subtilis strain and find that we are able to recapitulate the long deletions identified in previous experimental studies [2] and discover alternative combinations of deletions which have not yet been explored in vivo. MinGenome provides a pragmatic approach for constructing bioproduction chassis as genome reduction proceeds only up to the point that the minimized strain reaches growth and production yield and rate goals while alleviating the efforts to find the “true” minimal genome.

MinGenome: Top-down synthesis of genome minimized strains for bioproduction

255th National Meeting (2018)

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