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目的 构建重组人白细胞介素 10 (recombinanthumaninterleukin 10 ,rhIL 10 )融合蛋白的表达载体 ,并在大肠杆菌中表达。方法 应用RT PCR方法扩增IL 10基因 ,克隆PCR产物 ,构建PCRR○T7/NT TOPOR○ IL 10重组质粒 ,以AppiedBiosystems 370 0DNA分析仪进行分析。构建成功的重组质粒转化大肠杆菌BL2 1(DE3)pLysE细胞 ,经 12 %SDS PAGE鉴定融合表达蛋白。结果 PCRR○T7/NT TOPOR○ 质粒已载入rhIL 10基因 ,其序列与理论设计完全一致 ,表达质粒在BL2 1(DE3)pLysE中得到高效表达 ,产物主要以包涵体形式存在。结论 已成功构建重组PCRR○T7/NT TOPOR○ IL 10质粒载体 ,并在大肠杆菌BL2 1(DE3)pLysE细胞内高效表达
Objective To construct an expression vector for recombinant human interleukin 10 (rhIL 10) fusion protein and express it in E. coli. Methods The IL 10 gene was amplified by RT-PCR and the PCR product was cloned. The recombinant PCRR ○ T7 / NT TOPOR ○ IL 10 was constructed and analyzed by Appied Biosystems 370 0 DNA Analyzer. The constructed recombinant plasmids were transformed into E. coli BL21 (DE3) pLysE cells, and the expressed proteins were identified by 12% SDS PAGE. Results PCRR ○ T7 / NT TOPOR ○ plasmid was loaded with rhIL 10 gene. The sequence was consistent with the theoretical design. The expression plasmid was highly expressed in BL21 (DE3) pLysE, and the product was mainly in the form of inclusion body. Conclusion The recombinant PCRR ○ T7 / NT TOPOR ○ IL 10 plasmid vector has been successfully constructed and highly expressed in E. coli BL21 (DE3) pLysE cells