重组人白细胞介素-10在大肠杆菌中的表达及生物学活性分析

目的在大肠杆菌中表达重组人白细胞介素-10(Interleukin-10,IL-10),并检测其生物学活性。方法将IL-10基因重组到质粒pET11c中,转化BL21(DE),提取质粒,经酶切鉴定和测序分析;在25℃用低浓度的IPTG诱导表达,对包涵体IL-10稀释复性;经ELISA检测其含量,MC/9细胞增殖法检测其生物学活性。结果工程菌IL-10/pET11c/BL21诱导表达的目的蛋白以可溶性和包涵体两种形式存在,Westernblot鉴定证实为IL-10蛋白,两种形式的IL-10均具有一定的生物学活性。结论已成功地在大肠杆菌中表达了IL-10,为进一步纯化和制备IL-10的基因工程药物打下了基础。

Expression of Recombinant Human Interleukin-10 in E. coli and Biological Activity of Expressed Product

Objective To express recombinant human interleukin-10(IL-10) in E.coli and determine the biological activity of expressed product. Methods Insert IL-10 gene into expression vector pET11c and identify the recombinant pl asmid by restriction analysis and sequencing.Transform the recombinant plasmid t o E.coli for expression at 25℃ under induction of IPTG at a low concentrati on.Re-naturalize the expressed product,in a form of inclusion body,by dilution. Determine the content of target protein by ELISA and its biological activity by MC/9 cell proliferation method. Results The expressed product,existing in both soluble protein and inclusion body forms, was proved as IL-10 by Western blot and showed biological activity. Conclusion IL-10 was successfully expressed in E.coli.It laid a foundation of further purification and preparation of genetic engineering drug of IL-10.