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目的克隆hIL-22成熟链基因并在大肠杆菌表达体系中进行有效表达。方法采用反转录PCR以及巢式PCR的方法克隆得到hIL-22基因。将hIL-22基因装入PQE3.0载体构建重组载体hIL-22/PQE3.0,继而转化宿主工程菌株M15。经异丙基B-D硫代半乳糖苷诱导表达产生hIL-22/His重组蛋白并纯化。重组蛋白经电泳分析和Westernblot验证。结果成功地克隆和构建了hIL-22/PQE3.0载体,转化的工程菌经过诱导表达18kd的hIL-22成熟链,利用亲和层析得到了纯化的重组蛋白。结论成功获得hIL-22/His重组蛋白,为下一步研究人IL-22在肿瘤细胞中的作用奠定了物质基础。
Objective To clone the mature hIL-22 chain gene and express it in E.coli expression system. Methods The hIL-22 gene was cloned by reverse transcription PCR and nested PCR. The hIL-22 gene was inserted into PQE3.0 vector to construct recombinant vector hIL-22 / PQE3.0, which was transformed into host engineering strain M15. Induction of expression of hIL-22 / His with isopropyl B-D thiogalactoside resulted in the purification and purification of the recombinant protein. The recombinant protein was verified by electrophoresis and Western blot. Results The hIL-22 / PQE3.0 vector was successfully cloned and constructed. The transformed engineered bacteria induced the 18kd hIL-22 mature chain, and the purified recombinant protein was obtained by affinity chromatography. Conclusion The hIL-22 / His recombinant protein was successfully obtained, which laid the material foundation for the further study on the role of human IL-22 in tumor cells.