野生型人DNA polβ高表达中国仓鼠卵巢细胞系的建立及其生物学特征

脂质体转染法将野生型人DNA聚合酶beta(polβ)重组绿色荧光蛋白表达载体转染入中国仓鼠卵巢细胞系(CHO),经G418筛选得到稳定高表达人野生型polβ的CHO细胞.通过RT-PCR方法检测转染细胞polβmRNA的表达水平,流式细胞仪测细胞周期,软琼脂实验测细胞恶性增殖能力,6-TG实验测细胞自发突变率,以探讨转染细胞的生物学特性.结果显示,转染细胞polβ的mRNA的表达较空载体转染细胞、对照细胞增加,细胞周期多被阻滞在S期,细胞软琼脂集落形成率增高,自发突变率增加,表明polβ的过表达与细胞周期的分布、细胞恶性增殖程度、遗传稳定性相关.

Establishment of CHO Cell Line Stably Overexpressing Human Wild Type DNA Polymerase Beta Gene and Its Biological Characteristics

A recombined enhanced green fluorescent protein(GFP) vector pEGFP-AC3 carrying wide type DNA pol βgene was transfected into CHO cells by lipotransfection,and the stable transfectants were screened by G418.Stable overexpression of pol β in CHO was confirmed by RT-PCR,and its biological characteristics were analyzed from cell cycle,spontaneous mutation and cell soft agar growth.Expression level of pol β in the transfected cell increased significantly than the controls.The cell cycle distribution changed and most cells remained at S phage.And there was significant change in soft agar growths.Its spontaneous mutation rate was higher than that of the control cells.Overexpression of wild type pol β has some effect on the cell cycle distribution,malignant proliferation ability and genetic stability of cells.