白细胞介素-6基因的克隆、表达及纯化的研究

Interleukin-6 Gene Cloning, Expression and Purification

目的进行重组人白细胞介素-6(rhIL-6)的批量生产研究。方法将克隆的人IL-6基因置于pET30a载体的T7启动子下游,在宿主大肠杆菌中进行表达。结果工程菌表达量占菌体总蛋白的50%以上,纯品纯度大于95%,rhIL-6的分子量为21 000,等电点为6.7。用依赖IL-6的小鼠杂交瘤细胞系7TD1以MTT比色法测定表达蛋白生物活性表明,rhIL-6的ED50为0.35 ng/ml。结论上述结果表明rhIL-6已基本达到中试生产的要求。

Objective The main purpose of this paper is to study the batch production of recombinant human interleukin-6(rhIL-6). Methods The cloned rhIL-6 gene is under the control of T7 promoter of pET30a vector and expressed in E. coli. Results The ratio of expressed re- combinant protein to total cell protein is more than 50%. The rhIL-6 molecular weight is 21 000, isoelectric point is 6. 7. The purity of the rhIL-6 is more than 95%, and the activity of rhIL-6, determined by IL-6 dependent mice hybridoma cell line 7TD1 and MTT assay, is 0.35 ng/ml. Conclusions All results mentioned show that rhIL-6 meets the request of the middle scale production.