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Synthetic auxotrophs for stable and tunable maintenance of plasmid copy number

Author:
Kang, Chae Won, Lim, Hyun Gyu, Yang, Jina, Noh, Myung Hyun, Seo, Sang Woo, Jung, Gyoo Yeol
Source:
Metabolic engineering 2018 v.48 pp. 121-128
ISSN:
1096-7176
Subject:
Escherichia coli, antibiotics, auxotrophs, chromosomes, essential genes, gene expression, lycopene, plasmids, translation (genetics), value-added products
Abstract:
Although plasmid-based expression systems have advantages in multi-copy expression of genes, heterogeneity of plasmid copy number (PCN) in individual cells is inevitable even with the addition of antibiotics. Here, we developed a synthetic auxotrophic system for stable and tunable maintenance of the PCN in Escherichia coli without addition of antibiotics. This auxotroph expresses infA, one of the essential genes encoding a translation initiation factor, on a plasmid instead of on the chromosome. With this system, the gene expression was stably maintained for 40 generations with minimized cell-to-cell variation under antibiotic-free conditions. Moreover, varying the expression level of infA enabled us to rationally tune the PCN by more than 5.6-fold. This antibiotic-free PCN control system significantly improved the production of itaconic acid and lycopene compared to the conventional system based on antibiotics (2-fold). Collectively, the developed strategy could be a platform for the production of value-added products in antibiotic-free cultivation.
Agid:
5981107