羊抑制素α亚基N端1-33序列肽段多聚体的构建及其表达纯化  

Construction,Expression and Purification of Repeat Fragment Polymer of oInhibin α-Subunit N Terminal 1-33 Fragment

作　　者：黄运茂[1] 施振旦[1] 于迎春[1] 邵西兵[1] 

机构地区：[1]华南农业大学动物科学学院,广州510642

出　　处：《生物工程学报》2005年第2期311-314,共4页Chinese Journal of Biotechnology

基　　金：国家自然科学基金项目 (No .3 970 0 10 2 );广东省科技攻关项目 (No .2 0 0 3C2 0 2 0 5 )~~

摘　　要：为了提高短肽的免疫原性以制备短肽基因工程疫苗 ,将羊抑制素α亚基N端 1_33氨基酸残基片段的基因序列插入表达质粒pRSET A的BamHⅠ \SacⅠ位点之间构建重组质粒pR INH ,然后利用质粒中的一对同尾酶位点BamHⅠ \BglⅡ和下游的另一酶切位点HindⅢ ,经过简单的酶切后 ,将产物按各种组合连接 ,构建了抑制素串联 2至 6聚体基因。含 3至 6聚体基因的重组质粒pR 3INH、pR 4INH、pR 5INH和pR 6INH经IPTG诱导均能在大肠杆菌 (E .coli)BL2 1(DE3)中表达目的蛋白 ,分别占菌体蛋白的 6 %、6 %、7%和 8% ,且都以包涵体形式存在。结果说明 ,利用同尾酶切技术可以快速正确地构建短肽片段的串联多聚体重组质粒 ,为构建短肽半抗原的高免疫原性基因工程疫苗提供了新的思路。A cDNA sequence coding for ovine inhibin N terminal 1-33 AA residue fr agment (INH) was inserted between BamHⅠ\SacⅠ sites in plasmid pRS ET-A to gen erate plasmid pR-INH. By utilizing a pair of isocaudamer BamHⅠ and Bgl Ⅱ sites and another downstream HindⅢ site, following simple double diges tions and c ombination ligation of the resultant products, 2 to 6-repeat INH genes were con s tructed respectively. Each plamids containing 3 to 6 repeated INH fragment gene s, pR-3INH, pR-4INH, pR-5INH and pR-6INH, directed expression of the target prot eins in E.coli. BL21(DE3) under induction of ITPG, which respectively accoun ted for 6%, 6%, 7% and 8% of the total bacterial protein. The expressed target prot eins were all in the form of inclusion bodies. The above results implied that u tilization of isocaudamer restriction disgetion sites in expression plasmid is c apable of rapidly and correctly constructing repeat fragment polymer of short pe ptides, which may become a new method in construction of high immunogenic recomb inant vaccines of short peptides.

关 键 词：抑制素 同尾酶 串联多聚体 表达 

分 类 号：Q78[生物学—分子生物学]