Stable Maintenance of Plasmid in Continuous Culture of Yeast under Non-Selective Conditions

概要

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A recombinant yeast plasmid containing the gene for β-galactosidase was tested for stability in a host auxotrophic for leucine. Plasmid loss was studied at different dilution rates in continuous culture under selective as well as non-selective conditions. It was observed that the instability of the culture was higher at low dilution rates in selective medium, while the pattern was reversed when complex non-selective medium was used, with plasmid-containing cells competing effectively with plasmid-free cells at low dilution rates. This was attributed to a low residual yeast extract concentration in the medium at low dilution rates. Since yeast extract was the sole source of leucine, this limited the growth of plasmid-free cells, which were auxotrophic for leucine. Growth rate studies also indicated a competitive advantage of the plasmid-containing cells over the plasmid-free cells at low yeast extract concentrations in semi-defined medium. Using the above data, a modified continnuous culture was run using non-selective medium at a low dilution rate of 0.05 h^<-1>. This resulted in stable coexistence of plasmid-containing and plasmid-free cells and hence sustained expression of β-galactosidase at 〜330 OD_<420>l^<-1> h^<-1> throughout the period of cultivation (134h).
公益社団法人日本生物工学会の論文
2001-10-25