重组小鼠IL-33蛋白的原核克隆表达、纯化及生物学活性检测

Prokaryotic Cloning and Expression, Purification and Biological Activity Detection of Recombinant Mouse IL-33 Protein

目的:克隆鼠源白细胞介素33(murine interleukin-33)基因,构建大肠杆菌表达菌株,并对纯化后的蛋白进行生物活性的初步测定。方法:采用质粒抽提技术从大肠杆菌中提取质粒,构建p ET-32a-IL-33重组质粒;通过发酵罐培养技术获得蛋白,亲和层析方法纯化目的蛋白;ELISA检测纯化蛋白IL-33诱导产生TNF-α。结论:成功构建出鼠源IL-33的原核表达载体,成功表达出IL-33,IL-13能诱导TNF-α细胞因子的产生。

Objective:To clone murine interleukin-33(mIL-33)and express it in E.coli efficiently.Biological activity of the purified protein was measured.Methods:Plasmid were extracted from E.coli by plasmid extraction technique.The recombinant plasmid pET-32a-IL-33 was constructed by gene cloning.We used fermentor to culture the bacterica.IL-33 protein expression was induced by IPTG and purified by Ni-NTA affinity chromatography;the amount of TNF-αinducd by IL-33 was detected by ELISA.Conclusion:The whole-length cDNA of mouse IL-33 was successfully cloned and expressed,which laid a foundation for further studying on its biological function