EGFR胞内酪氨酸激酶结构域在人肿瘤细胞中的稳定表达  

EGFR tyrosine kinase domain stably expressed in human cancer cell line

作　　者：蔡志宇[1] 曲延征[1] 欧阳奇明[1] 黄晓晶[2] 陈泽辉[1] 

机构地区：[1]福建医科大学附属协和医院口腔颌面外科,福建福州350001 [2]福建医科大学附属口腔医院牙体牙髓科,福建福州350002

出　　处：《中国口腔颌面外科杂志》2006年第3期207-210,共4页China Journal of Oral and Maxillofacial Surgery

基　　金：国家留学基金;福建省自然科学基金(C0540009)

摘　　要：目的:探讨在人肿瘤细胞中稳定表达EGFR-TKD的方法。方法:采用磷酸钙共沉淀基因转染技术,将已构建的pEF-BOS/GST-EGFR-TKD质粒DNA与pBabe-puro质粒DNA共同导入体外培养的人非小细胞肺癌细胞系H1299中。加入嘌呤霉素,对转染的细胞加压筛选,以获得成功转染的细胞。免疫荧光法对重组质粒在H1299中的表达水平及定位进行检测。结果:经嘌呤霉素筛选,8d后对照组细胞全部死亡。转染组部分细胞存活,形成单克隆集落,经多次传代培养后,荧光显微镜下观察见多数集落的细胞胞质内表达GST-EGFR-TKD。结论:通过磷酸钙共沉淀法可成功转染H1299细胞,获得稳定表达EGFR-TKD的人肿瘤细胞。PURPOSE： This study is to find an approach for human cancer cell line to stably express EGFR- TKD. METHODS： H1299 （human non-small cell lung cancer cell line） cells were co-transfected with recombinant plasmid pEF-BOS/GST-EGFR-TKD and pBabe-puro by calcium phosphate transfection. Puromycin was added for selection. The expressing level and localization of protein products were detected by immuno-flourescence. RESULTS： 8 days after puromycin had been added, all cells in the control group died whereas a few cells in the transfection group survived and formed single colonies. After several passages, GST-EGFR-TKD could be detected in cell plasma in most of these colonies under flouresecence microscope. CONCLUSIONS： Human cancer cell stably expressing EGFR-TKD has been obtained by means of calcium phosphate transfection.

关 键 词：表皮生长因子受体 酪氨酸激酶结构域 人肿瘤细胞 体外转染 

分 类 号：R730.2[医药卫生—肿瘤]