| 乙型脑炎病毒SXBJ07株E基因的克隆及原核表达 |
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| 引用本文： | 王韦华,刘桂梅,张彦明.乙型脑炎病毒SXBJ07株E基因的克隆及原核表达[J].黑龙江畜牧兽医,2012(9):103-105. |
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| 作者姓名： | 王韦华 刘桂梅 张彦明 |
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| 作者单位： | 渭南职业技术学院;渭南市农产品食品执法监督支队;西北农林科技大学 |
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| 基金项目： | 国家自然科学基金项目(30771067) |
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| 摘 要： | 为构建乙型脑炎病毒(JEV)SXBJ07株E基因的原核表达载体,并在大肠杆菌中进行高效表达;根据JEV SXBJ07株基因组全序列设计1对特异性引物,RT-PCR扩增E基因全长;将目的基因插入pGEM-T连接载体,筛选出阳性重组质粒;将该质粒克隆至原核表达载体pET32α中,再转化入大肠杆菌BL21,经IPTG诱导表达后对其产物进行SDS-PAGE电泳分析和Western-blot检测。结果表明:扩增到了全长为1 500 bp的JEV SXBJ株E蛋白基因;重组质粒pET-32α-E构建成功;融合蛋白可以与乙脑阳性血清抗体特异性结合。说明在大肠杆菌中成功表达了JEV E蛋白。
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| 关 键 词： | 乙型脑炎病毒 E蛋白基因 基因克隆 原核表达载体 |
Cloning and expression in Escherichia coli of the Eprotein gene of Japanese encephalitis virus strain SXBJ07 |
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| WANG Wei-hua,LIU Gui-mei,ZHANG Yan-ming.Cloning and expression in Escherichia coli of the Eprotein gene of Japanese encephalitis virus strain SXBJ07[J].Heilongjiang Animal Science And veterinary Medicine,2012(9):103-105. |
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| Authors: | WANG Wei-hua LIU Gui-mei ZHANG Yan-ming |
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| Institution: | 1.Weinan Vocational and Technical College,Weinan 714000,China;2.Weinan Low Enforcement and Supervision Detachment of Agriculture Products and Food,Weinan 714000,China;3.Northwest A & F University,Yangling 712000,China) |
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| Abstract: | To construct the prokaryotic expression vector of the Eprotein gene of JEV strain SXBJ07 and obtain efficient expression in E.coli.A pair of specific primers was designed according to the full-length genomic sequences of JEV strain SXBJ07,and the E-protein gene was amplified by RT-PCR.The target gene was inserted into vector pGEM-T by ligation and the positive recombinant plasmid was selected.The recombinant plasmid was cloned into prokaryotic expression vector pET32alpha and transformed into E.coli BL21.After inducing with IPTG,the product was analyzed and determined using SDS-PAGE and Western-blot.The results showed that the E-protein gene with the length of 1 500 bp of JEV strain SXBJ was obtained by RT-PCR,the recombinant plasmid pET-32α-E was successfully constructed,and the fusion protein could specifically combined with the positive serum antibody of JEV.It indicates that the E-protein of JEV was successfully expressed in E.coli. |
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| Keywords: | Japanese encephalitis virus(JEV) E-protein gene gene cloning prokaryotic expression vector |
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