堆形艾美耳球虫巨噬细胞移动抑制因子的克隆和表达  

Cloning and expression of macrophage migration inhibitory factor from Eimeria acervulina

作　　者：阎晓菲[1,2] 韩红玉[1] 黄兵[1] 岳城[2] 赵其平[1] 姜连连[1] 董辉[1] 卞庆松[1] 张建哲[1] 

机构地区：[1]中国农业科学院上海兽医研究所／农业部动物寄生虫学重点开放实验室,上海200240 [2]新疆农业大学动物医学学院,新疆乌鲁木齐830052

出　　处：《中国预防兽医学报》2008年第11期856-860,共5页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家高技术研究发展计划项目(2006AA10A207-1);国家科技基础条件平台建设项目(2005DKA21104)

摘　　要：本研究扩增和克隆了堆形艾美耳球虫巨噬细胞移动抑制因子,并在大肠杆菌中表达出MIF重组蛋白。根据GenBank中的MIF mRNA序列,设计特异引物,采用PCR方法以堆形艾美耳球虫孢子化卵囊cDNA文库为模板扩增获得MIF基因,将MIF基因片段连接到原核表达载体pET-28a(+),构建重组表达质粒pET28a-MIF,并在大肠杆菌BL21(DE3)中诱导表达,对表达产物进行Western blot分析。结果从堆形艾美耳球虫孢子化卵囊cDNA文库中扩增出MIF基因片段,长度为709bp;经序列分析,扩增片段的核苷酸序列与GenBank中登录的MIF序列的同源性为99.5%;经SDS-PAGE检测表明重组质粒pET(28a)-MIF在大肠杆菌中以包涵体形式表达;Western blot分析证明堆形艾美耳球虫抗血清可与重组表达蛋白特异性结合。本研究结果为进一步研究该基因的功能奠定了基础。The macrophage migration inhibitory factor （MIF） of Eimeria acervulina were amplified by PCR from the cDNA library of E.acervulina spomlated oocysts and cloned into prokaryotic expression vector pET28a （＋）. The recombinant plasmid was transformed into BL21 （DE3） for expression and protein were identified by SDS-PAGE. The results showed that MIF gene fragment was 709 bp in length, which shared 99.5 % sequence identity with previously reported. SDS-PAGE showed the pET （28a）-MIF was expressed in the form of inclusion bodies. Western blot revealed that recombinant protein could be specifically recognized by antiserum of E.acervulina.

关 键 词：球虫 堆形艾美耳球虫 巨噬细胞移动抑制因子 克隆 表达 

分 类 号：S852.723[农业科学—基础兽医学]