[1]李荣,郑航,罗荣城.HER2基因的克隆及其在MCF-7细胞中的表达[J].南方医科大学学报,2005,(10):1264-1267.
 LI Rong,ZHENG Hang,LUO Rong-cheng.Cloning and expression of the HER2 gene in MCF-7 cells[J].,2005,(10):1264-1267.
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HER2基因的克隆及其在MCF-7细胞中的表达()
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《南方医科大学学报》[ISSN:/CN:]

卷:
期数:
2005年10期
页码:
1264-1267
栏目:
出版日期:
2005-10-01

文章信息/Info

Title:
Cloning and expression of the HER2 gene in MCF-7 cells
作者:
李荣 郑航 罗荣城
南方医科大学南方医院肿瘤科, 广东, 广州, 510515
Author(s):
LI Rong ZHENG Hang LUO Rong-cheng
Department of Oncology, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China
关键词:
人表皮生长因子受体雌激素受体HER2基因表达克隆转染乳腺肿瘤MCF-7细胞
Keywords:
human epidermal growth factor receptorestrogen receptorHER2gene expressioncloningtransfectionbreast neoplasmsMCF-7 cells
分类号:
R394.2;R737.9
摘要:
目的 克隆人表皮生长因子受体(HER2)基因,建立共表达HER2基因和雌激素受体(ER)基因的MCF-7细胞模型。方法 从高表达HER2的人乳腺癌细胞株SKBR-3中,用RT-PCR技术克隆HER2基因全长cDNA,将其插入载体pcDNA3.1中构建重组真核表达载体pcDNA3.1-HER2。测序鉴定后,利用阳离子脂质体介导将其转染ER阳性的乳腺癌MCF-7细胞,经G418筛选阳性克隆。用免疫细胞化学、流式细胞术及Westernblotting等方法检测HER2基因在MCF-7细胞中的稳定表达情况。结果 DNA测序证明,获得的HER2基因序列与GenBank中报道序列完全一致。将其转染MCF-7细胞,经流式细胞术、Westernblotting及免疫细胞化学等方法检测证实,转染细胞内有HER2基因高表达。结论 成功克隆HER2基因并获得共表达HER2基因和ER基因的MCF-7细胞株。
Abstract:
Objective To clone human epidermal growth factor receptor (HER2) gene and establish a MCF-7 cell line with stable co-expression of HER2 and estrogen receptor (ER). Methods The full-length HER2 gene fragment was amplified by RT-PCR from the total RNA extracted from human breast cancer cell line SKBR-3 which over-expressed HER2 protein. The gene fragment was then inserted into the vector pcDNA3.1 to construct the recombinant expression plasmid pcDNA3.1- HER2, which was identified by sequence analysis and transfected into ER-expressing MCF-7 cells via lipofectamine. The positive cell clones were obtained after G418 selection. The MCF-7 cells transfected with HER2-cDNA was assayed by immunocytochemistry, flow cytometry and Western blotting. Results DNA sequencing results showed that HER2 gene was exactly consistent with the sequence reported in GenBank. The MCF-7 cells tansfected with HER2 could highly express HER2 protein as shown by immunocytochemistry, flow cytometry and Western blotting. Conclusion HER2 gene has been cloned successfully and the MCF-7 cell line co-expressing HER2 and ER is obtained.

参考文献/References:

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备注/Memo

备注/Memo:
收稿日期:2005-5-12。
基金项目:广东省自然科学基金(037050)
作者简介:李荣(1974- ),男,在读博士研究生,主治医师,电话:020-61641656
通讯作者:罗荣城,电话:020-61641651,E-mail:center@nfhoc.com
更新日期/Last Update: 1900-01-01