| 用改进的重叠PCR引入血管内皮生长因子基因突变 |
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| 引用本文： | 白向阳,吕安国,吴文芳,孙静,牛瑞芳. 用改进的重叠PCR引入血管内皮生长因子基因突变[J]. 生物化学与生物物理进展, 2004, 31(2): 181-184 |
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| 作者姓名： | 白向阳 吕安国 吴文芳 孙静 牛瑞芳 |
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| 作者单位： | 1. 中国科学院沈阳应用生态研究所,沈阳,110016
2. 天津医科大学肿瘤医院中心实验室,天津,300060 |
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| 摘 要： | 血管内皮生长因子（vascular endothelial growth factor, VEGF）的PCR产物克隆于T载体上，经转化JM109感受态菌株后,随机挑取8个白斑菌落，混合后制成混合模板.采用3条引物，做两轮重叠PCR反应，获得了VEGF的突变基因，经PCR鉴定，酶切鉴定和测序分析表明所得基因为目的产物.实践证明这种突变方法简单快速，为下一步实验大量引入突变奠定了实验基础.
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| 关 键 词： | 点突变，血管内皮生长因子，重叠PCR |
| 收稿时间： | 2003-06-30 |
| 修稿时间： | 2003-08-28 |
Site-Directed Mutagenesis of Vascular Endothelial Growth Factor (VEGF) by An Improved Overlap PCR |
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| Affiliation: | Shenyang Institute of Applied Ecology, The Chinese Academy of Sciences, Shenyang 110016, China;Shenyang Institute of Applied Ecology, The Chinese Academy of Sciences, Shenyang 110016, China;Shenyang Institute of Applied Ecology, The Chinese Academy of Sciences, Shenyang 110016, China;Shenyang Institute of Applied Ecology, The Chinese Academy of Sciences, Shenyang 110016, China;Oncology Central Laboratory Cancer Hospital, Tianjin Medical University, Tianjin 300060, China |
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| Abstract: | The PCR production of vascular endothelial growth factor (VEGF) was subcloned into a T vector, and the recombined plasmids were then transformed into competent cell JM109. Eight white clones were selected randomly, and the recombined plasmids were extracted and mixed as template. Three selected primers were adapted and two rounds of PCR were performed to introduce mutations. The production of those were confirmed by enzyme digestion, PCR amplification and sequencing. The method is proved to be a simple,quick and easy way to introduce site directed mutagenesis, and make latter work easy to do. |
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| Keywords: | site-directed mutagenesis vascular endothelial grow factor overlap-PCR |
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