Application of Cross-Flow Filtration for High Density Cultivation and Regulation of trp Promoter

Abstract

A new fermentation system with continuous separation by cross-flow filtration using a ceramic filter was developed. By applying this system, high concentration cultivation of lactic acid bacteria and regulation of trp promoter for induction of a foreign gene by adjusting tryptophan concentration in a medium were carried out.

Streptococcus cremoris, Lactobacillus casei and Bifidobacterium longum were grown in the new fermentor up to high concentrations of 81, 49 and 54 g dry cell weight per liter, respectively. The cell productivities achieved by the fermentation system were 5 to 15 times as high as that of each corresponding conventional batch cultivation.

Escherichia coli harboring the recombinant plasmid containing β-galactosidase gene fused to trp promoter was cultivated in the fermentor with cross-flow filtration. In the initial stage of the cultivation, tryptophan concentration in a medium was kept at 170 µg ml⁻¹. The cross-flow filtration was started when optical density of cells at 570 nm became 10. Trytophan concentration dropped to low level within 30 min and an efficient expression of β-galactosidase gene was started. By this two stage cultivation, very high cell mass was achieved (60 g dry cell weight per liter) and amount of produced β-galactosidase was about 10% of total cellular protein.