| 表达HIV-1复合多表位基因的p815细胞稳定株的建立和评价 |
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| 引用本文： | 张宁,李英辉,赵亚,刘忠湘,毛张翔,黄豫晓,薛采芳.表达HIV-1复合多表位基因的p815细胞稳定株的建立和评价[J].科学技术与工程,2009,9(2). |
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| 作者姓名： | 张宁 李英辉 赵亚 刘忠湘 毛张翔 黄豫晓 薛采芳 |
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| 作者单位： | 第四军医大学病原生物学教研室,西安,710032 |
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| 摘 要： | 建立稳定表达HIV-1p24MEG复合多表位基因的p815细胞克隆.设计引物,以HIV-1标准株全长cDNA序列为模板,PCR扩增获得p24基因片段;合成改造后的多个表位基因MEG并且与p24片段相连接,克隆入pcDNA3.1(+).在阳离子聚合物作用下重组真核质粒转染的p815(H-2d)细胞, 以G418压力筛选,RT-PCR检测mRNA表达,间接免疫荧光检测蛋白表达.通过PCR获得了HIV-1 p24片段,获得了与多表位基因连接后的p24MEG融合基因,成功构建了p24MEG基因的重组真核表达载体pcDNA3.1(+)/p24MEG.转染p815细胞后, RT-PCR检测到融合蛋白mRNA表达,间接免疫荧光显示 p815细胞内有融合蛋白的表达.结论:建立了稳定表达HIV-1p24MEG融合蛋白的p815细胞克隆,为评价多表位抗原p24MEG在BALB/c小鼠体内诱导的细胞免疫应答奠定基础.
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| 关 键 词： | 复合多表位基因 稳定株 |
Construction and Screening the P815 Cell Clone Stably Expressing of HIV-1 Fusion Multi-epitope Gene |
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| ZHANG Ning,LI Ying-hui,ZHAO Ya,LIU Zhong-xiang,MAO Zhang-xiang,HUANG Yu-xiao,XUE Cai-fang.Construction and Screening the P815 Cell Clone Stably Expressing of HIV-1 Fusion Multi-epitope Gene[J].Science Technology and Engineering,2009,9(2). |
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| Authors: | ZHANG Ning LI Ying-hui ZHAO Ya LIU Zhong-xiang MAO Zhang-xiang HUANG Yu-xiao XUE Cai-fang |
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| Institution: | Department of Pathogenic Biology Fourth Military Medical University;Xi'an 710032;P.R.China |
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| Abstract: | The P815 cell clone stably expressing of HIV-1p24MEG fusion multi-epitope gene was constructed and screened.p24 gene was amplified by PCR,with the plasmid containing full-length of cDNA of HIV-1 standard strain as the template.The p24 gene and the gene coding cryptic epitopes by amino modified acid Substitution were inserted into pcDNA3.1(+),obtained the recombinant plasmid pcDNA3.1(+)/p24MEG.The P815(H-2d)cells were transfected with pcDNA3.1(+)/p24MEG by liposome.After screening by G418,positive cell clone... |
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| Keywords: | HIV-1 |
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