What genes constitute the fundamental building blocks of life? Surprisingly the identifies and functions of the absolute core set of essential genes required to sustain free-living self-replicating life remain unknown. To answer this question, we aim to employ novel methods to iteratively delete large and non-contiguous regions of the Escherichia coli genome to experimentally identify this core set of life-sustaining genes.
Previous work constructed a minimal ~530 kb synthetic Mycoplasma genome, 50 kb smaller than the ~580 kb M. genitalium genome. However, over 30% of the remaining genes are not functionally annotated. Taking advantage of the extensive characterization of the E. coli genome, we aim to use this as the model system to understand which genes and corresponding functions are required in a minimal self-replicating system. We plan to consolidate all permissive genetic deletions to construct a fully viable minimized E. coli strain, with the larger goal of generating the smallest E. coli genome ever created. This minimized strain will help to identify the core set of essential genes and corresponding functions required for life on earth, in addition to providing valuable insights into how genomes have evolved to increase cellular complexity and meet metabolic requirements.