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对新型降钙素基因工程菌 n CT/p GEX-2 T/E.coli BL2 1高表达发酵条件进行了优化研究。研究表明重组工程菌遗传性状稳定 ,采用 LB培养基、1 %接种量 ,当菌体浓度到达 A6 0 0 nm 0 .6时 ,添加诱导剂 IPTG至终浓度为 0 .l mmol/L,37℃培养 4 h,可使含新型降钙素的融合蛋白高效表达 ,融合蛋白占菌体总蛋白的 35%。2 0 L发酵罐中采用上述条件发酵培养 ,融合蛋白亦可高效表达 ( 32 % ) ,其菌体密度是摇瓶培养发酵的两倍。Western-blotting试验表明融合蛋白中含有新型降钙素 ,纯化制备的新型降钙素具有降钙活性
The optimization of fermentation conditions of high expression of nCT / p GEX-2 T / E.coli BL21 induced by a novel calcitonin gene engineering strain was optimized. The results showed that the genetic engineering of the recombined engineering bacteria was stable with LB medium and 1% inoculum. When the concentration reached A6 0 0 nm 0. 6, IPTG was added to the final concentration of 0.1 mmol / L and 37 ℃ After cultured for 4 h, the fusion protein containing novel calcitonin could be efficiently expressed, and the fusion protein accounted for 35% of the total bacterial protein. Fermentation culture was carried out in a 20 L fermenter under the above conditions, and the fusion protein was also highly expressed (32%). The cell density was twice that of shake flask culture. Western-blotting experiments showed that the fusion protein contains a new type of calcitonin, purified prepared new calcitonin with calcium lowering activity