转染hEGF基因的表皮细胞G418长期筛选后的表达  

The expression level of the transfected hEGF gene to cultured human epidermal cells

作　　者：董肇杨[1] 陈玉林[1] 戴方平[1] 郇京宁[1] 张素贞[1] 应康[1] 吕洛[1] 范清源[1] 方之扬[1] 

机构地区：[1]第二军医大学长海医院烧伤科

出　　处：《第二军医大学学报》1998年第S1期86-88,共3页Academic Journal of Second Military Medical University

基　　金：国家自然科学基金

摘　　要：目的：探讨外源性基因在转染表皮细胞后稳定表达情况。方法：选用构建的ｐＢＫ－Ｓｉｇｎａｌ－ＥＧＦ质粒，在脂质体ＬｉｐｏｆｅｃｔＡＭＩＮＥ的介导下，转染培养成６０％～７０％连片的人表皮细胞，然后用Ｇ４１８长时间筛选出稳定表达的细胞克隆。大量培养传代后，通过ＥＬＩＳＡ试剂盒检测培养上清中表皮细胞生长因子（ＥＧＦ）外源性基因稳定表达水平。结果：（１）构建的人表皮细胞生长因子（ｈＥＧＦ）质粒可成功转染到人表皮细胞中；（２）转基因表达在传４代后仍可存在；（３）ｈＥＧＦ表达水平６周内可在培养基中测出。Objective: To study the stable expression levels of cultured human epidermal cells transfected with a hEGFplasmid. Methods: The 60%-70% confluent cultured human epidermal cells were transfected a pBKSignalEGF plasmidconstruct mediated by lipofectAMINE. After screening of G418 successfully transfected, clone forming cells were picked and subcultured for several weeks. To detect hEGF expression levels and to determine long term transgenic stability, a specific ELISA was used. Results:(1) hEGF plasmid constructs were successfully introduced into cultured human epidermal cells. (2) Transgenic expression still persisted after four subcultures.(3) hEGF expression levels could be detected in the medium over a sixweek period. Conclusion: The stable expression of hEGF was obtained after gene transfected with the pBKSignalEGF plasmid mediated by lipofectAMINE.

关 键 词：基因转染 人表皮细胞生长因子 质粒 筛选 基因表达 

分 类 号：R644.03[医药卫生—外科学]