| H5N1禽流感病毒HA基因的重组腺病毒表达载体的构建 |
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| 引用本文： | 王青,胡建和,徐彦召,朱国坡,郑素玲.H5N1禽流感病毒HA基因的重组腺病毒表达载体的构建[J].河南农业科学,2010(7). |
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| 作者姓名： | 王青 胡建和 徐彦召 朱国坡 郑素玲 |
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| 作者单位： | 1. 河南科技学院,动物科学学院,河南,新乡,453003
2. 中国农业科学院,上海兽医研究所,上海,200241 |
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| 基金项目： | 河南省高校杰出科研人才创新工程项目 |
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| 摘 要： | 采用PCR方法扩增出禽流感病毒HA基因,将其定向插入pAdtrack-CMV腺病毒穿梭质粒中构建pAdtrack-H5,之后将pAdtrack-H5与腺病毒骨架质粒pAdeasy-1在基因工程菌BJ5183中进行同源重组,获得腺病毒质粒pAdeasy-H5,将pAdeasyd-H5经PacⅠ线性化后转染HEK293细胞株,包装出含有HA基因的腺病毒pAd-H5。结果表明:含有目的基因的腺病毒穿梭质粒pAdtrack-H5和含有目的基因的腺病毒质粒pAdeasy-H5经PCR、双酶切及核苷酸测序鉴定无误。包装成功的腺病毒pAd-H5经绿色荧光蛋白和RT-PCR分析证实,目的基因在该细胞中成功表达。
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| 关 键 词： | 腺病毒载体 禽流感病毒 H5N1亚型 HA基因 载体构建 |
Construction of A Recombinant Adenovirus Expression Vector Containing the HA Gene of H5N1 Avian Influenza Virus |
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| WANG Qing,HU Jian-he,XU Yan-zhao,ZHU Guo-po,ZHENG Su-ling.Construction of A Recombinant Adenovirus Expression Vector Containing the HA Gene of H5N1 Avian Influenza Virus[J].Journal of Henan Agricultural Sciences,2010(7). |
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| Authors: | WANG Qing HU Jian-he XU Yan-zhao ZHU Guo-po ZHENG Su-ling |
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| Abstract: | In the present work,the HA gene of avian influenza virus strain H5N1 was amplified from the plasmid pMD-19T-H5 by PCR and inserted into pAdtrack-CMV to construct a shuttle plasmid pAdtrack-H5.Then,the shuttle plasmid was transformed into BJ5183,which containing adenovirus backbone vector pAdeasy-1,to obtain recombinant adenovirus genome plasmid named as pAdeasy-H5.Both pAdtrack-H5 and pAdeasy-H5 were further identified by PCR,double-digestion and DNA sequencing.Then,pAdeasy-H5 was linearized with PacⅠ and transfected into HEK293 cells to form pAd-H5 viruses.GFP and RT-PCR were used to detect the expression of HA gene and the results showed HA pAdeasy-H5 was successfully expressed in HEK 293 cells. |
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| Keywords: | Recombinant adenovirus vector Avian influenza virus H5N1 HA gene Vector construction |
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