| 小鼠生殖细胞特异基因Oct4启动子片段克隆及报告载体构建 |
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| 引用本文： | 刘文强,曹晖,殷吉庆,云彦,华进联,雷安民.小鼠生殖细胞特异基因Oct4启动子片段克隆及报告载体构建[J].农业生物技术学报,2010,18(1):61-65. |
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| 作者姓名： | 刘文强 曹晖 殷吉庆 云彦 华进联 雷安民 |
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| 作者单位： | 西北农林科技大学动物医学院,陕西省干细胞工程技术研究中心,陕西省农业分子生物学重点实验室,杨凌,712100 |
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| 基金项目： | 留学回国人员科研启动项目，教育部重点科研项目，中国博士后科学基金特别资助，留学回国人员科研启动项目 |
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| 摘 要： | 本研究通过PCR方法从小鼠(Mus musculus)基因组中扩增出Oct4启动子的CR4区和CR1区,将其克隆到真核报告载体pEGFP-1,构建了携带Dct4启动子片段的小鼠生殖细胞特异性报告载体CR1,4-pEGFP-1.用该载体转染多种细胞,在荧光显微镜下观察绿色荧光蛋白(EGFP)在细胞中的表达情况,同时以半定量RT-PCR检测转染细胞中Oct4及其它生殖细胞特异性基因的转录情况.结果显示,GFP在小鼠胚胎干细胞、小鼠胚胎成纤维细胞(MEF)和C2C12细胞中均不表达,而在P19细胞和睾丸细胞中表达.RT-PCR结果显示,Oct4在P19、睾丸细胞和生殖干细胞中特异性转录.这表明所构建的绿色荧光蛋白报告载体只在生殖细胞和多能生殖干细胞中内表达,该质粒可作为示踪生殖细胞的工具,同时也可以作为生殖干细胞多能性状态的一个监测标记.
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| 关 键 词： | 生殖细胞 转染 |
Cloning of Mouse Germ Cell Tissue-specific Oct4 Promoter Fragment and Construction of Report Vector |
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| Liu Wenqiang Cao Hui Yin Jiqing Yun Yan Hua Jinlian Lei Anmin College of Veterinary Medicine,Shaanxi Stem Cell Engineering , Technology Research Center,Shaanxi Key Laboratory of Molecular Biology for Agriculture,Northwest Agriculture , Forestry University,Yangling.Cloning of Mouse Germ Cell Tissue-specific Oct4 Promoter Fragment and Construction of Report Vector[J].Journal of Agricultural Biotechnology,2010,18(1):61-65. |
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| Authors: | Liu Wenqiang Cao Hui Yin Jiqing Yun Yan Hua Jinlian Lei Anmin College of Veterinary Medicine Shaanxi Stem Cell Engineering Technology Research Center Shaanxi Key Laboratory of Molecular Biology for Agriculture Northwest Agriculture Forestry University Yangling |
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| Institution: | Liu Wenqiang Cao Hui Yin Jiqing Yun Yan Hua Jinlian Lei Anmin * College of Veterinary Medicine,Shaanxi Stem Cell Engineering , Technology Research Center,Shaanxi Key Laboratory of Molecular Biology for Agriculture,Northwest Agriculture , Forestry University,Yangling,712100 |
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| Abstract: | CR4 and CR1 fragments of mouse(Mus musculus) Oct4 promoter were cloned by PCR,inserted into eukaryotic report vector pEGFP-1 and reconstructed a mouse germ cell-specific report vector CR1,4-pEGFP-1.Recombinant plasmid was subsequently transfected into a variety of cells using Liposome 2000.Expression of the green fluorescent protein(EGFP) were detected under fluorescent microscopy.The expression of germ cell-specific gene were detected by semi-quantitative RT-PCR.The results showed that GFP was expressed in... |
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| Keywords: | Oct4 Oct4 Germ cells Transfection |
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