RV-G/LTB双基因融合真核表达质粒的构建及其在Vero细胞中的表达  

Construction of Eukaryotic Expression Vector for Fusion Gene RV-G/LTB and Its Expression in Vero Cells

作　　者：刘娟[1] 刘明远[1] 于录[1] 郭恒[1] 李扬[1] 赵丽晶[1] 孙树民[1] 李慧萍[1] 刘学[1] 王学林[1] 

机构地区：[1]人兽共患病研究教育部重点实验室吉林大学人兽共患病研究所,长春130062

出　　处：《中国生物制品学杂志》2011年第3期255-258,共4页Chinese Journal of Biologicals

摘　　要：目的构建RV-G/LTB双基因融合真核表达质粒,并在Vero细胞中表达。方法应用重叠延伸PCR方法,采用一定的linker序列(Gly4Ser)2将已克隆的狂犬病病毒糖蛋白(Rabies virus glycoprotein,RV-G)基因片段和大肠杆菌不耐热肠毒素B亚单位(E.coli heat-labile enterotoxin subunit B,LTB)基因片段进行融合,扩增融合基因G-linker-LTB,并插入真核表达质粒pVAX1中,构建重组真核表达质粒pVAX1-G-linker-LTB,转染Vero细胞,间接免疫荧光试验和Western blot法检测融合蛋白的表达。结果重组真核表达质粒pVAX1-G-linker-LTB经双酶切及测序鉴定证明构建正确,转染Vero细胞后,可检测到融合蛋白的表达。结论已成功构建了RV-G/LTB双基因融合真核表达质粒,并在Vero细胞中表达了融合蛋白。Objective To construct a eukaryotic expression vector for fusion gene RV-G / LTB and express in Vero cells.Methods The cloned gene fragments encoding rabies virus（RV） glycoprotein（G） and E.coli heat-labile enterotoxin subunit B（LTB） were fused by SOE-PCR using a linker sequence（Gly4Ser）2 and inserted into eukaryotic expression vector pVAX1.The constructed recombinant plasmid pVAX1-G-linker-LTB was transfected to Vero cells,and the expressed fusion protein was identified by indirect IFA and Western blot.Results Both restriction analysis and sequencing proved that recombinant plasmid pVAX1-G-linker-LTB was constructed correctly,and the expression of fusion protein was proved in transfected Vero cells.Conclusion A eukaryotic expression vector for fusion gene RV-G / LTB was successfully constructed and expressed in Vero cells.

关 键 词：狂犬病病毒 糖蛋白类 大肠杆菌不耐热肠毒素B 融合基因 VERO细胞 

分 类 号：Q78[生物学—分子生物学]