转移人端粒酶逆转录酶基因的实验研究

The experimental study of transferring the hTERT gene

目的 转移人端粒酶逆转录酶基因 (hTERT基因 ) ,建立其转移体系。方法 用脂质体转染的方法将逆转录病毒载体PLNC hTERT转入 ψ 2细胞 ,用产病毒的 ψ 2细胞克隆培养上清重复感染PA317细胞 ,建立产毒PA317/hTERT细胞 ;通过感染内皮细胞检测基因转移效果。结果 ψ 2细胞培养上清重复感染 ,使PA317细胞所产病毒滴度提高 2 0倍 ,得到高滴度的PA317/hTERT包装细胞系。用该细胞系产生的病毒上清感染内皮细胞成功。

Objective To establish the human telomerase reverse transcriptase (hTERT) gene viral transferring system. Methods By means of liposome mediation, a retroviral vector pLNC-hTERT carrying a selectable marker neomycin resistance genes (Neo r) and a target gene (hTERT) was transferred into the ectropic package cells ψ-2, and by using the supernatant of ψ-2 cells to infect the amphotropic producer clone PA317, established PA317/hTERT package cell line. In order to get higher-titer virus, the supernatants of ψ-2 infected PA317 repeatedly. Human endothelial cells were used to detect the effect of gene transfer. Results We established a PA317/hTERT package cell line which produced high-titer virus. The viral titer produced by PA317 cells was raised 20 times by repeated infection method, and with higher-titer virus the exogenous genes were successfully transferred into endothelial cell. Conclusion This method can be used to establish higher-titer viral transferring system.