| 小鼠FABP3基因全长cDNA的克隆、真核表达载体的构建及其在COS-7细胞中的表达 |
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| 引用本文： | 徐力致,李励芸,齐秋锋,闻娟,陈慧梅.小鼠FABP3基因全长cDNA的克隆、真核表达载体的构建及其在COS-7细胞中的表达[J].解放军医学杂志,2010,35(5). |
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| 作者姓名： | 徐力致 李励芸 齐秋锋 闻娟 陈慧梅 |
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| 作者单位： | 南京大学医学院医学遗传学教研室,江苏省医学分子技术重点实验室,南京,210093 |
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| 基金项目： | 国家自然科学基金(30800546);;教育部博士点基金(200802841008) |
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| 摘 要： | 目的克隆小鼠FABP3基因的全长cDNA,构建其真核表达载体,并于COS-7细胞内表达。方法利用PCR技术扩增小鼠FABP3基因的开放阅读框序列,并在3′末端连接24bp的Flag标签,克隆至真核表达载体pcDNA3.1中,构建表达质粒pcD-NA3.1-FABP3和pcDNA3.1-FABP3-Flag,并行PCR、双酶切及测序鉴定。将构建的重组质粒通过脂质体转染COS-7细胞,分别采用RT-PCR和免疫荧光法检测其FABP3mRNA及蛋白的表达情况。结果所构建的pcDNA3.1-FABP3和pcDNA3.1-FABP3-Flag重组载体可酶切出418bp的FABP3 cDNA片段和442bp的FABP3-FlagcDNA片段,经测序证实正确。转染后的COS-7细胞经RT-PCR可扩增出278bp的目的片段,且免疫荧光分析可见特异性的FABP3及Flag蛋白表达。结论构建了FABP3基因真核表达载体pcDNA3.1-FABP3和pcDNA3.1-FABP3-Flag,并于COS-7细胞中成功转录表达,为后续研究奠定了基础。
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| 关 键 词： | 脂肪酸结合蛋白质类 克隆 分子 遗传载体 COS细胞 |
Cloning of full-length cDNA of mouse FABP3 gene,construction of eukaryotic expression vector and expression of the vector in COS-7 cells |
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| XU Li-zhi,LI Li-yun,QI Qiu-feng,WEN Juan,CHEN Hui-mei.Cloning of full-length cDNA of mouse FABP3 gene,construction of eukaryotic expression vector and expression of the vector in COS-7 cells[J].Medical Journal of Chinese People's Liberation Army,2010,35(5). |
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| Authors: | XU Li-zhi LI Li-yun QI Qiu-feng WEN Juan CHEN Hui-mei |
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| Abstract: | Objective It had been found in previous gene expression microarray study that the gene of fatty acid binding protein 3 (FABP3) might contribute to the renal damage of patients with obesity related glomerulopathy. For further exploring the effect of FABP3 gene on podocyte lesion related obesity, the objective of present study was to construct the eukaryotic expression vector containing FABP3 gene, and to detect the expression of the vector in COS-7 cells. Methods The specific primers were designed according ... |
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| Keywords: | fatty acid-binding proteins cloning molecular genetic vectors COS cells |
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