摘要
目的获得ＩＬ－１８全长ｃＤＮＡ序列，研究ＩＬ－１８在大肠杆菌中的表达。方法利用ＲＴ－ＰＣＲ方法从人外周血单个核细胞中扩增出人白细胞介素（ｈＩＬ－１８）的ｃＤＮＡ，以Ｓａｎｇｅｒ双脱氧法测序。进而构建了含ｒｅｃＡ启动子的表达载体ＰＭＯＮｈＩＬ－１８，并在大肠杆菌ＪＭ１０１中表达，制备包涵体，并用ＳＤＳ－ＰＡＧＥ分析表达产物。结果ＲＴ－ＰＣＲ扩增出约５００ｂｐ片段，ＳＤＳ－ＰＡＧＥ示一约１８ｋＤ的蛋白，与预期分子量相符，表达量占菌体蛋白的２０％。结论ＩＬ－１８在ｒｅｃＡ启动子下获得了高效表达，ＩＬ－１８位于细胞包涵体内。
Objective To obtain the cDNA of human interleukin-18(hIL-18) for the study of its expression.Methods The human IL-18 cDNA was obtained from human PBMC by RT-PCR. The cDNA was sequenced and cloned into the expression vector containing rec A promoter. The recombinant protein was expressed in E. coli strain JM101 and the inclusion body was separated and analyzed by SDS-PAGE. Results A 500bp DNA fragment was obtained by RT-PCR. The sequence of the cDNA was consistent with the published hIL-18 cDNA. After induction with nalidixic acid, SDS-PAGE showed a new protein with the molecular mass of 18kD. The yield of hIL-18 was about 20% of the total cellular protein. Conclusion hIL-18 was highly expressed under the rec A promoter and the expressed hIL-18 was mainly in the inclusion body.
出处
《徐州医学院学报》
CAS
1998年第6期441-444,共4页
Acta Academiae Medicinae Xuzhou
