绵羊CD58基因的克隆与原核表达  

Cloning and prokaryotic expression of sheep CD58 gene

作　　者：芦晓立[1,2] 颜新敏[2] 吴国华[2] 叶奕优[3] 李健[2] 朱海霞[2] 王建科[2] 赵志荀[1,2] 崔力凡[2] 朱彩珠[2] 张强[2] 

机构地区：[1]甘肃农业大学动物医学院,甘肃兰州730070 [2]中国农业科学院兰州兽医研究所,家畜病原生物学国家重点实验室,农业部畜禽病毒学重点开放实验室,甘肃兰州730046 [3]深圳出入境检验检疫局动植物检验检疫技术中心,广东深圳518001

出　　处：《甘肃农业大学学报》2010年第5期11-14,28,共5页Journal of Gansu Agricultural University

基　　金：甘肃省重大科技专项(092NKDA032);国家科技支撑计划项目(2006BAD06A11;2006BAD06A17)

摘　　要：通过对猪、绵羊CD58mRNA序列比对,设计出绵羊CD58的特异性引物,应用反转录PCR技术从绵羊淋巴细胞mRNA中扩增出绵羊CD58基因,将该基因克隆到原核表达载体pGEX4-1中进行原核表达.结果表明:克隆出的绵羊CD58cDNA长807bp,其中包含了开放式阅读框;该基因在大肠杆菌中进行表达,SDS-PAGE电泳在53ku处能观察到特异表达的融合蛋白,产物通过Western-blot检测证实,可以被CD58单克隆抗体识别.CD58 plays an important role in mammal immune system.In this study,according to the porcine and sheep CD58 sequences,reference to the porcine CD58 mRNA primers,apair of primers was designed and used to clone sheep CD58 gene in lymphocyte mRNA by RT-PCR method.Then,this cDNA was ligated into pGEX-4T-1 and transformed to E.coli BL21cells.The results showed that an 807bp fragment of sheep CD58 cDNA with an ORF of 762bp was obtained.The recombinant transformant was induced with IPTG,SDS-PAGE showed the specific expression of the fusion protein was observed in 53ku and the expressed product could be recognized by CD58 monoclonal antibody in Western-blot.This research will help us to develop new adjuvant and immune-regulated drugs,and laid a solid foundation for further study on the structure of CD58 molecule and the mechanism of CD2-CD58 complex triggering immune system.

关 键 词：绵羊 CD58基因 基因克隆 原核表达 

分 类 号：S852.4[农业科学—基础兽医学]