过表达ERCC1蛋白细胞系的建立

Establishment of over-expressing ERCC1 protein cell line

关键词:  ERCC1  构建  转染  基因表达  细胞系

Key words:  ERCC1  construction  transfection  gene expression  cell line

摘要:

建立过表达ERCC1蛋白的细胞系，为遗传毒性化合物的研究提供模型细胞系，并有可能为具有DNA加合作用化合物提供新的毒理学筛选方法。［方法］ 应用PCR和DNA重组技术将核苷酸切除修复交叉互补组1(Excision repair cross complementation group1, ERCC1)基因与pT7Blue载体相连接，然后构建真核细胞重组表达质粒pcDNA3.1/Zeo(+)-ERCC1，并导入293细胞。经Zeocin抗性筛选后获得转染ERCC1基因的细胞克隆。细胞增殖后以Western-blot检测目的蛋白的表达。［结果］ 获得稳定转染ERCC1基因的细胞系，并在细胞中检测出有ERCC1蛋白的过表达。［结论］ 用基因转染法成功建立了过表达ERCC1蛋白的ERCC1-293细胞系

Abstract:

To establish an over-expressing ERCC1 protein model cell line ERCC1-293 to be used for study of genotoxic chemicals. ［Methods］ Excision repair cross complementation group1(ERCC1) gene was transferred to pT7Blue vector with polymerase chain reaction (PCR) and DNA recombinant techniques. Recombinant express plasmid pcDNA3.1/Zeo(+)-ERCC1 was constructed and introduced into 293 cells. The ERCC1 transfected cell clones were obtained with Zeocin selection. The expression of ERCC1 protein was detected by Western-blot after the cells being proliferated. ［Results］ The model cell line transfected with ERCC1 was gained, which could over-express the ERCC1 protein in cells. ［Conclusion］ An over-expressing ERCC1 protein model cell line is established successfully by gene transfection method.