EFFECT OF GROWTH-RATE ON STABILITY AND GENE-EXPRESSION OF RECOMBINANTPLASMIDS DURING CONTINUOUS AND HIGH CELL-DENSITY CULTIVATION OF ESCHERICHIA-COLI TG1

Continuous and fed-batch cultures of recombinant Escherichia coli TG1 were carried out in order to study plasmid stability and recombinant p roduct formation at different specific growth rates. The aprotinin=bet a-galactosidase gene (Ap=lacZ) was placed under the control of two dif ferent promoter/repressor systems, the P(Lac)/lacI (pPLac8) and the la mbdaP(L)/cI(ts857) (pPL6) system. The chemically (0.5 mM IPTG) induced gene expression exhibited higher product activity and plasmid stabili ty than the thermally (40-degrees-C) induced expression. In fed-batch cultivations with the more stable E. coli TG1(pPLac8) a special feedin g strategy allowed bacterial growth with a constant growth rate mu for several hours up to high cell densities. The cloned gene product acti vity was noticeably effected by the specific growth rate and the cell density at the moment of induction. In particular, the enzyme activiti es passed a pronounced maximum value in dependence of the set growth r ate. The results indicate that fed-batch cultivation strategies are we ll suited to produce recombinant gene products.