| 人Wnt10b基因真核表达载体的构建及表达鉴定 |
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| 引用本文： | 纪影畅,蔡湘娜,林常敏,李宇.人Wnt10b基因真核表达载体的构建及表达鉴定[J].汕头大学医学院学报,2011,24(4):196-199,190. |
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| 作者姓名： | 纪影畅 蔡湘娜 林常敏 李宇 |
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| 作者单位： | 1. 汕头大学医学院第一附属医院整形外科,广东汕头,515041
2. 汕头大学医学院组织学与胚胎学教研室,广东汕头,515041 |
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| 基金项目： | 国家自然科学基金资助项目(81071577); 广东省自然科学基金资助项目(9151008901000195); 广东省科技计划资助项目(2009B030801320); 广东省医学科研基金资助项目(A2009433) |
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| 摘 要： | 目的：利用基因工程技术构建人Wnt10b基因的pEGFP-N1-Wnt10b真核表达载体,观察其在COS-7细胞中的表达。方法：以pUC19-Wnt10b为模板,PCR扩增人Wnt10b基因的cDNA编码区序列,扩增产物和pEGFP-N1载体双酶切,T4 DNA连接酶连接,构建pEGFP-N1-Wnt10b,酶切和测序鉴定该载体;LipofectamineTM2000将该载体转染入COS-7中,Western blot和免疫细胞化学检测COS-7 Wnt10b蛋白表达。结果：PCR扩增出的人Wnt10b基因cDNA正确克隆到pEGFP-N1载体,成功构建了pEGFP-N1-Wnt10b;将其转染入COS-7,COS-7 Wnt10b蛋白表达明显增高。结论：成功构建了人pEGFP-N1-Wnt10b,这为进一步研究Wnt10b基因功能奠定了前期基础。
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| 关 键 词： | Wnt10b基因 载体 COS-7细胞 基因表达 |
Construction and Identification of Eukaryotic Expression Vector Bearing Human Wnt10b Gene |
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| JI Ying-chang
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CAI Xiang-na
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LIN Chang-min
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LI Yu.Construction and Identification of Eukaryotic Expression Vector Bearing Human Wnt10b Gene[J].Journal of Shantou University Medical College,2011,24(4):196-199,190. |
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| Authors: | JI Ying-chang CAI Xiang-na LIN Chang-min LI Yu |
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| Affiliation: | JI Ying-chang1,CAI Xiang-na1,LIN Chang-min2,LI Yu1(Department of Plastic Surgery,The First Affiliated Hospital,Shantou University Medical College,Shantou 515041,China) |
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| Abstract: | Objective:To construct human Wnt10b gene eukaryotic expression vector,and to observe its expression in transient transfeted COS-7 cell line.Methods:cDNA fragment was amplified from pUC19-Wnt10b by PCR and cloned into eukaryotic expression vector pEGFP-N1.The positive clone was confirmed by enzyme digestion and sequencing.The recombinant plasmid was transiently transfected into COS-7 cell line with LipofectamineTM2000.The expression of Wnt10b gene was detected by Western blot and immunocytochemistry.Results:... |
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| Keywords: | Wnt10b gene vector COS-7 cell gene expression |
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