| shRNA-resistant Gankyrin质粒的构建及鉴定 |
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| 引用本文： | 赵青.shRNA-resistant Gankyrin质粒的构建及鉴定[J].科学技术与工程,2013,13(12):3238-3241. |
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| 作者姓名： | 赵青 |
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| 作者单位： | 解放军总医院 国家生物医学分析中心 |
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| 基金项目： | The National Natural Science Foundation of China (General Program, Key Program, Major Research Plan) |
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| 摘 要： | gankyrin是近年来发现的一种癌基因,研究表明其表达产物Gankyrin与肿瘤发生密切相关。实验室前期工作发现Gankyrin表达水平与临床乳腺癌转移存在正相关,并可能通过多个环节影响乳腺癌的转移。本研究利用PCR介导的定点突变技术,构建了shRNA-resistant Gankyrin抵抗型质粒。经酶切鉴定及DNA测序证实序列完全正确,抵抗型质粒构建成功;经Western Blot检测,抵抗型质粒在HEK293T细胞中正确表达,为进一步研究Gankyrin在乳腺癌转移中的作用及其分子机制提供了重要的实验材料。
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| 关 键 词： | Gankyrin 定点突变 抵抗型质粒 RNA干扰回复 |
| 收稿时间： | 2012/12/21 0:00:00 |
| 修稿时间： | 1/21/2013 7:13:45 PM |
Construction and expression of shRNA-resistant Gankyrin plasmid |
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| zhaoqing.Construction and expression of shRNA-resistant Gankyrin plasmid[J].Science Technology and Engineering,2013,13(12):3238-3241. |
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| Authors: | zhaoqing |
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| Institution: | 2(The General Hospital of Chinese People’s Liberation Army1,Beijing 100853,P.R.China; National Center of Biomedical Analysis2,Beijing 100850,P.R.China) |
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| Abstract: | Gankyrin, an oncoprotein, has been reported to be associated with tumorigenesis. Previous study in our laboratory showed that Gankyrin was highly expressed in human breast cancer tissues than in adjacent normal tissues and affected the breast cancer metastasis in many stages. In this study, a shRNA-resistant Gankyrin plasmid is constructed by PCR induced site-directed mutagenesis. Identification by restriction endonuclease digestion and sequencing analysis indicated that the resistant plasmid was constructed successfully. Moreover, the expression of shRNA-resistant Gankyrin plasmid was detected in HEK293T cells by Western blot. This study provides an important tool for the further research of Gankyrin functions and its molecular mechanism in Breast Cancer. |
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| Keywords: | Gankyrin site-directed mutagenesis resistant plasmid RNAi rescue |
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