Instability of trp operon recombinant plasmid in E. coli

Abstract

The stability of ColEl-trp recombinant plasmid was studied in Escherichia coli mutant strains. MV12/pVH5 trp$^+$ strains were stable during the cultivation up to stationary phase regardless of the composition of medium. In death phase, trp$^-$ cells were generated at a frequency of 1\% and one of 4 trp$^-$ cells had three modified plasmids and the rest had no plasmid. When MV12/pVH5 trpR$^+$ strain was cultivated in chemostat culture without tryptophan, stability of the plasmid was maintained at 100\% until 50 generation times. In glucose-limited chemostat culture, trp$^-$ cells began to appear after about 30 generation times and they were enriched in the culture medium due to the higher growth rate than trp$^+$ cells. In MV12/pVH5 tna$^-$ strains and in the cultivation with indoleacrylic acid ($30\mu$g/ml), the stability was decreased by about 10\%. In these conditions, trp$^-$ cells showed the heterogeneous profile of plasmids whose sizes were smaller and/or 2-3 times larger than original ColEl-trp plasmid. However, in MV12/pVH5 trpR$^-$ strains, the stability of the plasmid was decreased by about 95\% because trp$^-$ cells were enriched in the culture medium due to the higher growth rate than trp$^+$ cells, and the trp$^-$ cells were lossed their entire plasmids. In trp$^-$ cells which appeared in death phase, the interconversion between trp$^+$ and trp$^-$ cells was detected and it is believed that this event was related to the expression of trp operon.