以端粒酶活性为作用靶点筛选抗肿瘤药的一种快速简易方法

A Quickly and Simplified Method for Screening Anticancer Drug in Telomerase Activity as a Target

利用ＰＣＲ扩增端粒重复序列，通过电泳和银染色测定扩增产物，以电泳胶中６ｂｐ梯状条带确定端粒酶活性。温度３０℃，反应时间为６０ｍｉｎ时可以使酶提液与未知化合物作用后还保持着酶的活性。从电泳的上样量及不同浓度细胞酶提液的结果表明，最低上样量１０μｌ和细胞酶提液０５μｇ时，能得到清晰６ｂｐ的梯状条带。ＲＮａｓｅ０．０５μｇ对端粒酶活性显示抑制作用，而对ＴａｑＤＮＡ酶没有影响。

Telomerase activity was measured by the polymerase chain reaction(PCR) based telomeric repeat amplification protocol(TRAP). For analysis of the TRAP assay, the PCR products were separated by the gel electrophoresis. The 6bp DNA ladder was showed by the silver staining. Telomerase activity was observed at the temperature of 30℃ and in reaction time of 60 min. The results obtained varying PCR products quantities and telomerase extract from cell showed that distinct 6bp DNA ladder could be got at 10 μl PCR product quantity and 0.5 μg telomerase extract from tumor cell. Telomerase activity was inhibited at the concentration of 0.05 μg RNase, and no effect occurred for the Taq DNA polymerase.