| 鹅细小病毒延边株VP1基因真核表达质粒构建及其在Vero细胞中的表达 |
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| 引用本文： | 赵文婧,鲁承,杜秋明,高建伟,申峰勇,张颖,李闯,袁娜,房靖添.鹅细小病毒延边株VP1基因真核表达质粒构建及其在Vero细胞中的表达[J].畜牧与兽医,2012,44(2):14-17. |
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| 作者姓名： | 赵文婧 鲁承 杜秋明 高建伟 申峰勇 张颖 李闯 袁娜 房靖添 |
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| 作者单位： | 1. 延边大学农学院动物医学系,吉林延吉,133000
2. 延边州动物疫病预防控制中心,吉林延吉,133000 |
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| 摘 要： | 根据GenBank已发表的鹅细小病毒(GPV)B株基因序列,设计并合成1对含有Xho I和BamH I酶切位点的特异性引物,以提取的GPV基因组DNA为模板,经PCR扩增得到了1 163 bp的目的片段,并将该片段克隆至pMD-18T simple载体中,再将其亚克隆至真核表达载体pVAX1中,构建重组表达质粒pVAX1-VP1。经PCR鉴定、酶切分析和序列测定比较,证实了重组质粒的正确性。通过脂质体法将pVAX1-VP1转染Vero细胞,经间接荧光抗体检测,在Vero细胞表面可见特异性荧光。经RT-PCR扩增得1 163 bp的目的片段。该研究为GPV核酸疫苗的研制奠定了基础。
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| 关 键 词： | 鹅细小病毒 VP1基因 真核表达 |
Construction of eukaryotic expression plasmid for VP1 gene of goose parvovirus isolated from Yanbian and its expression in Vero cells |
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| ZHAO Wen-jing
,
LU Cheng
,
DU Qiu-ming
,
GAO Jian-wei
,
SHEN Feng-yong
,
ZHANG Ying
,
LI Chuang
,
YUAN Na
,
FANG Jing-tian.Construction of eukaryotic expression plasmid for VP1 gene of goose parvovirus isolated from Yanbian and its expression in Vero cells[J].Animal Husbandry & Veterinary Medicine,2012,44(2):14-17. |
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| Authors: | ZHAO Wen-jing LU Cheng DU Qiu-ming GAO Jian-wei SHEN Feng-yong ZHANG Ying LI Chuang YUAN Na FANG Jing-tian |
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| Institution: | 1 (1.Veterinary Department,Agricultural College of Yanbian University,Yanji 133000,China; 2.Yanbian Korean Autonomous Prefecture Animal Disease Control Centre,Yanji 133000,China) |
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| Abstract: | According to the published gene sequence of goose parvovirus(GPV) strain B in GenBank,a pair of specific primers with Xho I and BamH I restriction enzyme sites were designed and synthesized.The VP1 gene fragment of GPV was amplified by PCR and cloned into pMD18-T simple vector.The fragment was subcloned to eukaryotic expression vector pVAX1 and the recombinant plasmid pVAX1-VP1 was identified by PCR,double restriction enzyme analysis and sequence analysis.Then the recombinant pVAX1-VP1 was transfected to Vero cell with Lipofectamine 2000.The transfected cells were detected by the indirect fluorescent antibody.The specific fluorescence could be observed on the cell surface.The 1 163 bp DNA fragment was amplified from RNA extracted from the Vero cell transfected with pVAX1-VP1 by RT-PCR. |
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| Keywords: | goose parvovirus VP1 gene eukaryotic expression |
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