Abstract
In the future, entire genomes tailored to specific functions and environments could be designed using computational tools. However, computational tools for genome design are currently scarce. Here we present algorithms that enable the use of design-simulate-test cycles for genome design, using genome minimisation as a proof-of-concept. Minimal genomes are ideal for this purpose as they have a simple functional assay whether the cell replicates or not. We used the first (and currently only published) whole-cell model for the bacterium Mycoplasma genitalium. Our computational design-simulate-test cycles discovered novel in silico minimal genomes which, if biologically correct, predict in vivo genomes smaller than JCVI-Syn3.0; a bacterium with, currently, the smallest genome that can be grown in pure culture. In the process, we identified 10 low essential genes and produced evidence for at least two Mycoplasma genitalium in silico minimal genomes. This work brings combined computational and laboratory genome engineering a step closer.
Highlights
In the future, entire genomes tailored to specific functions and environments could be designed using computational tools
We produced two genome design algorithms (Minesweeper and the Guess/Add/Mate Algorithm (GAMA)) which use the M. genitalium whole-cell model to generate minimal genome designs. Using these computational tools we found functional in silico minimal genomes which, if biologically correct, produce in vivo predictions between 33 and 52 genes smaller than the most recent predictions for a reduced Mycoplasma genome of 413 genes[17]
We created two genome design algorithms (Minesweeper and GAMA) that used computational design-simulate-test cycles to produce in silico M. genitalium minimal genomes (Minesweeper_256 and GAMA_237, 36% and 41% in silico reductions, respectively)
Summary
Entire genomes tailored to specific functions and environments could be designed using computational tools. 1234567890():,; For genome engineering and design, minimal genomes are currently the best proof-of-concept[1] These are reduced genomes containing only genes essential for life, provided there is a rich growth medium and no external stressors[1,2]. The greatest progress to date includes: JCVI-Syn3.0, a 50% gene reduction of Mycoplasma mycoides[2]; several strains of Escherichia coli reduced by 38.93 and 35%4 of their base pairs in vivo; an E. coli gene reduction of 77.6% in Saccharomyces cerevisiae[5]; and two 36% gene reductions of Bacillus subtilis[6] These efforts began with prescriptive design, gene selection using existing knowledge, or based on laboratory testing of individual genes, followed by iterative development. This process is timeconsuming and expensive due to the limitations of current techniques and unexpected cell death. Some non-essential genes become essential when a functionally-equivalent gene is removed, leaving a single pathway to a metabolite
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