人白细胞介素13重组蛋白表达载体的构建及其在大肠埃希菌的表达

Construction of Recombinant Human Interleukin-13 Protein Expression Vector and Its Expression in Escherichia coli

目的 构建人白细胞介素 13(rh IL- 13)重组蛋白表达载体 ,热诱导 rh IL- 13在 E.coli DH5 α表达。方法 经巢式 RT- PCR从非粘附贴壁生长的 PBMC活化细胞中克隆 h IL- 13成熟蛋白编码基因 ,随后构建 rh IL13- PBV重组蛋白表达质粒 ,热诱导表达重组蛋白。结果 在 h IL13- PBV/ DH5 α工程菌热诱导全菌蛋白中检测到相对分子质量大小约为 10× 10 3的 h IL- 13重组蛋白 (rh IL- 13蛋白 )。结论 rh IL- 13的表达成功 ,为今后进一步研究 h IL-13的生物学功能 ,探讨其临床应用价值奠定了一定的物质基础。

OBJECTIVE To construct recombinant human interleukin 13 protein(rhIL 13) expression vector with its expression in E.coli.METHODS The target cDNA fragment encoding the mature human interleukin 13 (hIL 13) protein was amplified from the total RNA,which were come from activated non adherent human PBMC(peripheral blood mononuclear cells), by nested reverse transcriptase polymerase chain reaction (RT PCR).Then it was cloned into the expression plasmid vector pBV220 containing a thermo inducible promoter P RP L.The constructed expression vector pBV220 containing the hIL 13 cDNA target fragment was named as hIL13 PBV and the engineering bacteria,the E.coli DH5α tranformed with hIL13 PBV was named as hIL13 PBV/DH5α.RESULTS After heat induction for the hIL13 PBV/5α by shifting the culture temperature from 30℃ to 42℃,the recombinant hIL 13(rhIL 13) protein was expressed with a relative molecular mass about 10×10 3 proved by SDS PAGE.CONCLUSIONS Successfully expression of rhIL 13 was propitious to the later researches on the biologic function of IL 13, and providing a substance basis for the clinical application of IL 13 in the future.