| DEV-NP基因酵母双杂交诱饵载体的构建及自激活检测 |
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| 引用本文： | 张荷,代淑燕,毛君婷,王开功,周碧君,文明.DEV-NP基因酵母双杂交诱饵载体的构建及自激活检测[J].生物技术通报,2011(3). |
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| 作者姓名： | 张荷 代淑燕 毛君婷 王开功 周碧君 文明 |
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| 作者单位： | 1. 贵州大学动物科学学院,贵阳,550025
2. 贵州大学动物科学学院,贵阳,550025;贵州省动物疫病研究室,贵阳,550025 |
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| 摘 要： | 为构建鸭肠炎病毒(DEV)核衣壳蛋白(NP)基因酵母双杂交诱饵载体,采用PCR技术扩增出DEV-NP基因,克隆至酵母双杂交诱饵质粒pGBKT7中,以PCR检测、限制性酶切和序列测定等方法进行鉴定;然后采用PEG/LiA.法将阳性质粒pGBKT7-NP转化酵母Y2HGold,在不同营养缺失型培养基进行自激活检测,结果显示,经PCR检测和EcoR I/BamH I双酶切,可见到与预期相一致的目的片段;测序表明该片段含DEV NP基因全部序列;转化Y2HGold酵母菌后,在SD/-Trp/X-a-Gal 固体培养基上长出蓝色菌落,而在SD/-Trp/X-a-GaUAbA固体培养基上未见有菌落生长.无自激活作用的诱饵载体pGBKT7NP的成功构建,为DEV NP宿主结合蛋白的筛选莫定了基础.
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| 关 键 词： | 鸭肠炎病毒核衣壳蛋白 诱饵载体 酵母双杂交 自激活作用 |
Construction of a Bait Vector for DEV NP Gene and Evaluation of Autoactivation in Yeast Two-hybrid System |
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| Zhang He,Dai Shuyan,Mao Junting,Wang Kaigong,Zhou Bijun,Wen Ming.Construction of a Bait Vector for DEV NP Gene and Evaluation of Autoactivation in Yeast Two-hybrid System[J].Biotechnology Bulletin,2011(3). |
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| Authors: | Zhang He Dai Shuyan Mao Junting Wang Kaigong Zhou Bijun Wen Ming |
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| Affiliation: | Zhang He1 Dai Shuyan1 Mao Junting1 Wang Kaigong1,2 Zhou Bijun1,2 Wen Ming1,2(1College of Animal Science,Guizhou University,Guiyang 550025,2Institute of Animal Disease,Guiyang Province,Guiyang 550025) |
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| Abstract: | To construct a bait vector of nucleocapsid for duck enteritis virus and to evaluate autoactivation in yeast two-hybrid system.Full-length of NP gene was amplified by polymerase chain reaction(PCR) and confirmed by sequencing.A fragment of the gene was then subcloned into MCS of pGBKT7 vector to construct the bait vector according to the reading frame.The reconstructed bait vector pGBKT7-NP was transformed into the yeast strain Y2HGold by PEG/LiAc method and its self-activation was tested by the phenotype as... |
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| Keywords: | NP of DEV Bait vector Yeast two-hybrid system Autonomous reporter activation |
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