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《天津医科大学学报》[ISSN:1006-8147/CN:12-1259/R]

卷:

20卷

期数:

2014年06期

页码:

429-432

栏目:

基础医学

出版日期:

2014-11-20

文章信息/Info

Title:

Construction of myocyte enhancer factor 2A eukaryotic expression plasmid and effects on cell proliferation in breast cancer cell line MCF7

文章编号:

1006-8147（2014）06-0429-04

作者:

伦淑敏

(天津医科大学肿瘤医院肿瘤研究所,国家肿瘤临床医学研究中心,天津市肿瘤防治重点实验室 ,天津 300060)

Author(s):

LUN Shu-min

(Tianjin Medical University Cancer Institute and HospitalNational Clinical Research Center of Cancer, Ministry of Education Key Laboratory of Cancer Prevention and TherapyKey Laboratory of Cancer Prevention and TherapyTianjin 300060 China)

关键词:

肌细胞增强因子2A; 乳腺癌; 质粒; 增殖

Keywords:

myocyte enhancer factor 2A; breast cancer; plasmid; proliferation

分类号:

R737.9

DOI:

-

文献标志码:

A

摘要:

目的：构建MEF2A基因真核表达重组质粒，并转染乳腺癌细胞株，观察其对细胞增殖能力的影响。方法：实时定量聚合酶链反应（RT-QPCR）检测MCF-7、BT474及MDA-MB-231等3种乳腺癌细胞中MEF2A mRNA的表达。采用全基因合成法合成MEF2A编码区序列，克隆入pcDNA3.1（+）-HA载体，构建重组质粒pcDNA3.1-MEF2A-HA。重组质粒转染乳腺癌细胞，采用Western blot及RT-QPCR检测MEF2A的表达效率。细胞计数实验检测细胞的增殖能力。结果：3株乳腺癌细胞中，MDA-MB-231细胞中MEF2A mRNA表达水平较低。其基因测序结果显示与预期片段大小一致，表明MEF2A重组质粒构建成功。将MEF2A重组质粒瞬时转染MDA-MB-231细胞后， MEF2A表达质粒组与对照组比较，其MEF2A表达升高，细胞增殖能力增强。结论：成功构建MEF2A过表达重组质粒，在MDA-MB-231中过表达MEF2A促进细胞的增殖能力。

Abstract:

Objective: To construct eukaryotic expression plasmid of pcDNA3.1-MEF2A and examine its effect on the proliferation of breast cancer cell line. Methods: The MEF2A mRNA levels were detected in MCF7, BT474 and MDA-MB-231 cells, respectively by real-time quantitative PCR. The full-length coding sequence of MEF2A was amplified by standard RT-QPCR. The amplified。MEF2A was cloned into pcDA3.1 vector. The eukaryotic expression plasmid pcDNA3.1-MEF2A-HA was transfected into breast cancer cells. RT-QPCR and western blot were performed to measure the expression level of MEF2A. Cell counting was performed to evaluate the effects of MEF2A expression on cell proliferation. Results: The MEF2A mRNA expression level of MDA-MB-231 was low in MDA-MB-231 cells. Western blot and RT-QPCR indicated that transfection of MEF2A expression vector into MDA-MB-231 cells resulted in increased mRNA/protein levels of MEF2A. The forced MEF2A expression significantly enhanced cell proliferation of MDA-MB-231. Conclusion: The eukaryotic expression plasmid of pcDNA3.1-MEF2A-HA is constructed successfully. Over-expression of MEF2A enhances proliferation ability of MDA-MB-231cells.



 

参考文献/References:

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备注/Memo

备注/Memo:

作者简介 伦淑敏（1988-），女，硕士在读，研究方向：乳腺癌相关基因的转录调控机制；E-mail：lunshuminvs@163.com。

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更新日期/Last Update: 2014-11-20