pRL-hTNF/JM103工程菌发酵和rhTNFα表达的研究

Fermentation Process for pRL-hTNF/JM103 Engineering Bacterial Strain and Expression Condition of rhTNFα

为便于下游中试规模生产，采用温度敏感型启动子PRPL构建rhTNFα工程菌，并对影响工程菌发酵培养和表达的因素进行了初步研究。结果显示，采用RTB培养基，50℃热水快速升温，诱导培养4～4．5h，湿菌体收获率和表达量均达到较高水平；50L发酵罐连续发酵3批，14000r／min连续离心，菌体收获率湿重达16．gg／L培养物，rhTNFα表达量占菌体总蛋白的10.5％，活性达1．35×107U／mg蛋白。.

After pRL-rhTNF/JM013 engineering bacterial strain was constructed usingtemperature-sensitive promotor PRPL, the factors influencing the fermentation of it and theexpression of rhTNF α were preliminarily studied. The result showed that, by using RTBmedium, the recovery and expression rates of the strain reached high levels after being heatedwith water (50℃ ) and inducing at 42℃ for 4~4. 5h. Three lots of the engineering productswere separated by continuous centrifugation (14000r/min). The average recovery rate of rhTNF α reached 16. 9g/L cultrue, and the activity of it reached 1. 35 × 107U/mg protein, and10. 5% of total somatic protein was expressed.