EFFECTIVE PRODUCTION OF BIOLOGICAL SUBSTANCES BY HIGH CELL-DENSITY CULTIVATION

High concentration cultivation is effective to improve the productivity of biological substances in fermentation. Especially in the cultivation of gene-engineered microorganisms and animal cells, high concentration cultivation may give the most effective productivity. To attain high cell concentration, cntrolled feeding of nutrients is very important. Therefore, we developed an on-line glucose and an on-line organic acids monitoring and controlling systems. By using the systems, we could improve bakers yeast cultivation and lactic acid fermentation. By applying the on-line glucose controlling system for the culture of recombinant yeast, on-off control of the gene expression and the effective production of heterologous proteins with SUC and PGK expression vectors could be attained. The gene expression from Escherichia coli trp promoter was also well derepressed by by controlling glucose concentration. We also studied several problems with high cell density culture of recombinant microorganisms with such as E. coli runaway replication plasmid, trp, pho gene expression systems and yeast GAL expression vector. We could obtain suitable culture conditions for them and achieve the overproduction of cloned gene product. We also developed a nobel cell immobilization method with alginate and urethane polymer to achieve high cell density culture of animal cell. By this immobilization method, we could culture animal cells for at least 2 months with high cell density of more than 10(7) cells/cm3-gel.