| 共表达猪流行性腹泻病毒S1与GM-CSF蛋白重组腺病毒的构建 |
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| 引用本文： | 赵攀登,董建国,邓同炜,陈益,徐耀辉,蒋增海,卢建洲,王晓晔,王岩.共表达猪流行性腹泻病毒S1与GM-CSF蛋白重组腺病毒的构建[J].动物医学进展,2016(6). |
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| 作者姓名： | 赵攀登 董建国 邓同炜 陈益 徐耀辉 蒋增海 卢建洲 王晓晔 王岩 |
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| 作者单位： | 1. 河南牧业经济学院 动物医学院,河南郑州,450046;2. 信阳农林学院 牧医工程学院,河南信阳,464000;3. 广西大学 动物科学技术学院,广西南宁,530004 |
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| 摘 要： | 为构建表达猪流行性腹泻病毒(PEDV)S1与GM-CSF融合基因的重组腺病毒,通过FMDV的2A基因序列做为Linker将密码子优化的S1基因和粒细胞巨噬细胞集落刺激因子(GM-CSF)基因连接。构建重组穿梭质粒pShuttle-GM-CSF 2A-S1(m),并将其电转化至含有腺病毒骨架载体pAdEasy-1的大肠埃希菌BJ5183感受态细胞中,通过同源重组制备重组腺病毒质粒pAd-GMCSF2A-S1(m)。重组腺病毒质粒经PacI线性化后转染HEK-293A细胞,获得重组腺病毒rAd-GMCSF2A-S1。间接免疫荧光和Western blot结果显示,S1和GMCSF蛋白在重组腺病毒感染的HEK-293A细胞中获得表达。该重组腺病毒的制备为PEDV重组活载体疫苗的研究奠定了基础。
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| 关 键 词： | 猪流行性腹泻病毒 S1 基因 粒细胞巨噬细胞集落刺激因子 重组腺病毒 |
Construction of Recombinant Adenovirus Co-expressing PEDV S1 and GM-CSF Proteins |
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| Abstract: | In order to construct the recombinant adenovirus expressing PEDV S1 and GM-CSF genes,the co-don optimized S1 gene was linked with GM-CSF gene by FMDV 2A gene.Recombinant shuttle plasmid pShuttle-GMCSF2A-S1(m)was constructed and transformed into Escherichia coli BJ5183 competent cells containing adenovirus backbone vector pAdEasy-1 to produce pAd-GMCSF2A-S1 (m)by homologous re-combination.The recombinant adenovirus plasmid was linearized by PacI and was transfected into HEK-293A cells.The recombinant adenovirus rAd-GMCSF2A-S1 was rescued and identified by indirect immuno-fluorescence and Western blot assay.The results showed that S1 and GM-CSF proteins were expressed in HEK-293A cells infected with rAd-GMCSF2A-S1.This study laid a foundation for the development of re-combinant live vector vaccine against PEDV. |
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| Keywords: | Porcine epidemic diarrhea virus S1 gene GM-CSF recombinant adenovirus |
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