[3S02e] 05. Genome Editing Technology and its Applications - Part 3

5:25 PM - 5:45 PM JST(8:25 AM - 8:45 AM UTC)

[3GET-16] E. coli 0.5: Generating Escherichia coli controlled by a half-sized genome

○Takahito Mukai¹, Masayuki Su’etsugu¹ (1. Rikkyo Univ. (Japan))

Keywords:

genome synthesis

In the era of generative biology, genome synthesis might outperform genome editing. Here, we introduce our platform (GIGI) to generate and implement a genome instance in E. coli. To demonstrate the utility, a prototype minimal genome of E. coli was assembled and fleshed out to finally establish E. coli controlled by a half-sized genome. GIGI consists of an integrated development environment (IDE) and a runtime environment (RTE). The genome IDE supported the cell-free assembly of 200-300 kb plasmids and the in vivo fusion of these chromosomes into a single 1.7 Mb plasmid. The cellular RTE employes host cells installed with a genome self-digestion system in the presence or absence of the RecA recombinase. In the absence, the prototype genome never complemented the programmed elimination of the host genome. On the other hand, varieties of chimeric strains were developed via RecA-mediated homologous recombination between the guest and host genomes. Among them, REGE-244 has the smallest genome (2.44 Mb). Thus, the RTE was useful to test and debug a prototype genome, albeit in a rough manner. To update the RTE, a new host strain is prepared for the restriction enzyme-mediated zombification of E. coli (REZE). REZE is achieved by the programmed digestion of the host genome and elimination of the survivors. We aim to directly flesh out the genome skeleton in zombies cells via massive patching of the genome until the emergence of living cells.