| 蛋白表达抑制稳定株MDA—MB-231-Tudor—SNI的筛选和鉴定 |
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| 引用本文： | 王保亚,刘欣,高星杰,朱梦瑜,杨振霞,赵秀娟,杨洁.蛋白表达抑制稳定株MDA—MB-231-Tudor—SNI的筛选和鉴定[J].国外医学：分子生物学分册,2010(5):401-405. |
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| 作者姓名： | 王保亚 刘欣 高星杰 朱梦瑜 杨振霞 赵秀娟 杨洁 |
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| 作者单位： | [1]天津医科大学免疫学教研室,天津市300070 [2]国家教育部免疫微环境与疫病重点实验室,天津市300070 [3]天津市细胞与分子免疫学重点实验室,天津市300070 |
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| 基金项目： | 国家高新技术研究发展计划（863计划）（No.2007AA022115）,国家自然科学基金（No.30670441,30970562,30670802,30811130394,30911130165,30970582）,国家重点基础研究发展规划（973计划）（No.2009CB918903）,天津市科委国际合作项目（N0.07JCZDJC07300）,国家自然科学重大研究计划培育项目（No.90919032）,国家教育部高等学校博士学科点专项科研基金（No.20091202110001）,天津市教委重点项目（No.2008ZD01） |
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| 摘 要： | 目的采用针对人类Tudor—SN基因的RNA干扰（RNAi）技术建立Tudor—SN表达抑制的乳腺癌MDA—MB-231稳定细胞株。方法利用脂质体转染方法将pGenesil—shRNA—hTudor—SNI质粒转染进入MDA—MB-231细胞，经G418筛选出稳定株，通过RT—PCR检测Tudor—SN的mRNA水平，再以Western印迹技术鉴定Tudor—SN蛋白表达情况并计算蛋白表达抑制率。结果与对照组相比，以G418筛选的MDA—MB-231-Tudor—SNI稳定株Tudor—SN的mRNA水平降低（P〈0．01，n=3），蛋白表达水平明显下调（P〈0．01，n=3），蛋白表达抑制率达78．12％。结论成功筛选并鉴定人类Tudor—SN蛋白表达抑制MDA—MB-231稳定株，有助于深入研究Tudor—SN蛋白在乳腺癌中的作用。
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| 关 键 词： | 人类Tudor—SN蛋白 RNA干扰 细胞筛选 稳定株 |
MDA-MB-231-Tudor-SNI Cell Line |
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| Authors: | WANG Baoya LIU Xin GAO Xingjie ZHU Mengyu YANG Zhengxia ZHAO Xiujuan YANG Jie |
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| Institution: | ( Department of Immunology, Tianjin Medical University, Tianjin, 300070, China Key Laboratory of Immunology Microenuironment and Disease, Ministry of Education of China, Tianjin, 300070, China Tianjin Key Laboratory of Cellular and Molecular Immunology, Tianjin, 300070, China) |
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| Abstract: | Objective To transfect the constructed plasmid expressing down-regulation of Hu- man Tudor-SN Gene into MDA-MB-231 cells and screen the cell lines of stable interference human Tudor-SN gene. Methods Recombinant vector pGenesil-shRNA-hTudor-SNI was transfected into MDA-MB-231 cells using Lipofectamine-2000. The cell strains for stable and down-expression of Tudor-SN protein with the medium containing G418 sulfate were selected. The expression level of Tudor-SN was then detected by RT-PCR and western blotting assay. Results The hTudor-SN mRNA and protein expression levels in stable transfectant screened with G418 were down-regulated as com- pared to those in negative control group （P 〈0. 01, n = 3） . The inhibition rate of Tudor-SN protein expression was up to 78.12%. Conclusion The stable MDA-MB-231-Tudor-SNI cell lines were screened and examined successfully. The results are useful for research on the function of Tudor-SN protein on breast cancer. |
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| Keywords: | human Tudor-SN protein RNAi cell screening stable transfectant |
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