PRODUCTION OF INSECTICIDAL PROTEIN OF BACILLUS-THURINGIENSIS BY CULTIVATION OF RECOMBINANT ESCHERICHIA-COLI

Cultures of recombinant Escherichia coli JM103 harboring the 135 k dalton (Da) insecticidal protein (IP) gene from Bacillus thuringiensis subsp. aizawai strain IPL7 were investigated to obtain high cell density and a large amount of 135kDa IP. In test tube cultures of recombinant E. coli JM103, soybean hydrolysate was apparently effective in improving 135kDa IP production. However, when fed-batch culture was carried out with soybean hydrolysate, a large amount of acetate accumulated in the broth and acetate was evidently the main inhibitor of cell growth and 135kDa IP production. In recombinant E. coli JM101, JM105 and JM109 producing 135kDa IP, the addition of soybean hydrolysate also increased 135kDa IP production, but it stimulated acetate accumulation as well. In order to remove acetate from the broth, cultivation with cross-flow filtration was employed. The final cell concentration of recombinant E. coli JM103 was 125 g/dm3, and 135kDa IP was produced at a concentration of 6.6 g/dm3.