带His-tag的乙肝病毒融合抗原在毕赤氏酵母中的表达  

Expression of HBV Fusion Antigen SpreS1 with Polyhistidine-tags in Pichia pastoris

作　　者：胡翔华[1] 刘南松[1] 吕红[1] 袁汉英[1] 李育阳[1] 

机构地区：[1]复旦大学生命科学学院遗传学研究所遗传工程国家重点实验室,上海200433

出　　处：《复旦学报（自然科学版）》2005年第4期560-565,共6页Journal of Fudan University：Natural Science

基　　金：国家高技术研究发展计划资助项目(Z18-01-02-02)

摘　　要：乙肝病毒融合抗原SpreS1(简称SS1抗原)较S抗原有更强的免疫原性,可望有更好的临床应用前景.为了便于表达产物的纯化,利用化学合成的寡聚核苷酸链和PCR扩增,在SS1基因的5’端和3’端分别融合了6His-EK和6His的编码序列,将其重组质粒pPIC3.5k-6His-EK-SS1、pPIC3.5k-SS1-6His转化毕赤氏酵母菌GS115,筛选鉴定得到的重组酵母工程菌进行诱导培养,然后采用镍离子柱(Ni-NTABasin)纯化表达产物,纯化的蛋白样品经ELISA效价测定、SDS-PAGE和WesternBlot分析,结果表明融合抗原基因6His-EK-SS1和SS1-6His在毕赤氏酵母中均能表达,纯化的产物仍有免疫原性,所用的纯化方法简便有效.Hepatitis B virus surface recombine antigen SpreS1 has higher immunity and better clinic apply foreground than S antigen. SpreS1 has successfully expressed intracellular in Pichia pastoris, for the purpose of facilitated purification, 6-Histidine-tag and 6-Histidine-EK-tag were separately introduced into 3＇ and 5＇ terminus of SpreS1-encoding DNA and the fusion genes were subcloned into the expression vector pPIC3.5k. The resultant recombinant plasmids pPIC3.5k-6His-EK-SpreSland pPIC3.5k-SpreS1 6His were separately transformed into Pichia pastoris GSll5. The effects of 6-His-tag and the fusion position on the expression of SpreS1 and the immunity of recombinant fusion SpreS1 were studied. Both pPIC3.5k-6His-EK-SpreSland pPIC3.5k-SpreS1-6His can express in Pichia pastoris, and the purified 6His-EK-SpreS1 exhibited strong immunity under ELISA and Western Blot analysis.

关 键 词：乙肝重组抗原SpreS1 毕赤氏酵母菌 镍离子柱 融合抗原 表达产物 毕赤氏酵母 乙肝病毒 HIS-TAG SDS-PAGE Western 

分 类 号：Q943.2[生物学—植物学] Q786