|Stabilization of an E. coli plasmid by a mini-F fragment of DNA.
J. Industrial. Microbiol. 2: 323-328. 1988.
H. Yukawa, Y. Kurusu, M. Shimazu, H. Yamagata and M. Terasawa.
|In an Escherichia coli K-12 strain (trpA trpE tnaA) cultured in LB broth without selective pressure, a pBR322 derivative containing the gene for tryptophan synthase (pBR322-trpBA) was found to be unstable.After 70 cell-number doublings, only 50% of the host cells retained the gene for amipicillin resistance (Apr). Insertion of the mini-F fragment of F factor DNA into this plasmid could effectively reduce the plasmid loss. Partial repression of the tryptophan promotor-operator by 3-indoleacrylic acid further decreased the stability of the pBR322-trpBA but not that of themini-F inserted plasmid (pBR322F-trpBA). The vector pBR322F-trpBA could be maintained at high copy number in the culture after 100 generations of growth; the culture was able to overproduce tryptophan synthate in the presence of 3-indoleacrylic acid. L-Tryptophan was produced from indole and L-serin using an E. coli host transformed with pBR322F-trpBA DNA. After 8 h of incubation, the expression level was approximately 180 g/l.|