| 猪繁殖与呼吸综合征病毒河北地方株ORF6基因的克隆及原核表达 |
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| 引用本文： | 王娇,孙继国,陈赛娟,刘涛,王坤,赵泽坤.猪繁殖与呼吸综合征病毒河北地方株ORF6基因的克隆及原核表达[J].中国动物检疫,2009,26(4):34-36. |
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| 作者姓名： | 王娇 孙继国 陈赛娟 刘涛 王坤 赵泽坤 |
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| 作者单位： | 1. 河北农业大学动物科技学院,河北保定,071001
2. 商务部流通产业促进中心,北京,100070 |
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| 基金项目： | 河北省重大技术创新项目 |
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| 摘 要： | 从河北沧州分离到一株疑似猪繁殖与呼吸综合征病毒,接种Marc-145细胞,经4代盲传,出现细胞病变,经鉴定为PRRSV,命名为HB-3(cz)株。根据GenBank公布的PRRSV JXA1株ORF6基因的核苷酸序列,设计并合成一对特异性引物(P1/P2),用RT-PCR方法扩增完整ORF6基因,将扩增产物连接到pGM-T载体并转化克隆菌,阳性重组质粒PGM-M进行序列测定与分析。后将克隆质粒PGM-M双酶切后连接原核表达载体pET-32a(+),在Rosseta-DE3中成功获得表达,经Western-blotting分析表明,表达蛋白与阳性血清发生特异性反应。
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| 关 键 词： | 猪繁殖与呼吸综合征病毒 河北分离株 ORF6基因 原核表达 |
Preliminary studies of prokaryotic expression of ORF6 gene of porcine reproductive and respiratory syndrome virus HB-3(cz) strain |
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| WANG Jiao,SUN Jiguo,CHEN Saijuan,LIU Tao,WANG Kun,ZHAO Zekun.Preliminary studies of prokaryotic expression of ORF6 gene of porcine reproductive and respiratory syndrome virus HB-3(cz) strain[J].China Journal Of Animal Quarantine,2009,26(4):34-36. |
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| Authors: | WANG Jiao SUN Jiguo CHEN Saijuan LIU Tao WANG Kun ZHAO Zekun |
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| Institution: | 1 (1.College of Animal Science and Technologe,Agricultural University of Hebei,Baoding 071001,China; 2. Circulation Industry Promotion Center of the Ministry of Commerce,Beijing100070,China) |
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| Abstract: | The full-length of ORF6 gene encoding the envelope glycoprotein (GP6) of PRRSV HB-3(cz) strain isolated from Cangzhou HeBei was amplified by RT-PCR with a pair of specific primers (p1/ p2) designed according to the relevant sequence of PRRSV JXA1 strain, and then cloned into plasmid pGM-T,the recombinant plasmid designated as pGM-M was subjected to sequencing and analysis. After identification by restriction endonclease digestion,the gene was subcloned into prokaryotic expression vector pET-32 a (+). The recombinant fusion proteins pET-M were highly expressed in Escherichia coli BL21. Western-blotting showed that the recombinant protein could react with the porcine polyclonal antibody against PRRSV. |
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| Keywords: | Porcine reproductive and respiratory syndrome virus HB-3(cz) strain ORF6 gene prokaryotic expression |
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