穿透支原体P35蛋白对细胞因子的诱生作用

被引：4

作者：

朱翠明

吴移谋

万艳平

余敏君

机构：

[1] 南华大学微生物学与免疫学教研室

[2] 南华大学微生物学与免疫学教研室衡阳

[3] 衡阳

来源：

中华微生物学和免疫学杂志 | 2004年 / 09期

基金：

湖南省自然科学基金;

关键词：

穿透支原体; P35; 细胞因子;

D O I：

暂无

中图分类号：

R375 [支原体属];

学科分类号：

100103 ; 100705 ;

摘要：

目的　构建pQE31/p35原核细胞表达载体 ,研究穿透支原体 (Mpe)相对分子质量 (Mr)为 35× 10 3 蛋白 (P35 )诱生巨噬细胞 (MΦ)分泌TNF α、IL 1β、IL 6等细胞因子 (CKs)的作用。方法　PCR扩增p35全长 10 0 2bp基因片段 ,克隆至pQE31原核细胞表达载体 ,用PCR介导的突变将p35基因中两个“TGA”终止密码子定点突变为“TGG” ,使其在大肠杆菌 (E .coli)中编码表达色氨酸。IPTG诱导pQE31/p35在E .coli中表达目的蛋白rP35 ,用Ni NTASpin亲和纯化 ,Westernblot鉴定表达产物。用酶联免疫吸附试验 (ELISA)检测P35刺激小鼠MΦ分泌TNF α、IL 1β和IL 6的量。结果　PCR扩增出约 1kb的DNA片段 ,克隆至pQE31载体 ,筛选到阳性克隆pQE31/p35 ,p35基因中两个“TGA”终止密码子成功突变成“TGG”。IPTG诱导pQE31/p35在E .coli中表达出Mr 约 37× 10 3 的蛋白质 ,Westernblot鉴定其为rP35。ELISA结果表明 ,P35能刺激小鼠MΦ分泌大量的TNF α、IL 1β和IL 6。结论　P35能诱导MΦ分泌TNF α、IL 1β、IL 6等细胞因子

引用

页码：37 / 40

页数：4

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