乙基亚硝基脲对人白血病细胞HPRT基因的影响

Effect of ethylnitrosourea on HPRT gene in human promyelocytic leukemia cells

目的 探讨乙基亚硝基脲 (ENU)诱导人次黄嘌呤鸟嘌呤磷酸核糖基转移酶 (HPRT)基因突变的分子图谱和发生机制。方法 采用单细胞克隆培养、双向筛选计数、多重聚合酶链反应扩增和电泳分析等方法。结果 随着ENU剂量的增加 ,细胞接种存活率非常显著下降 (10 0～ 2 0 0 μg/ml剂量组 )、突变频率显著升高 (12 5～ 2 0 0 0 μg/ml剂量组 )。自发突变没有全部基因外显子缺失型 ,只有 7 7%检测出单个外显子缺失 ;而ENU诱发突变中却有 79 7%显示缺失改变 ,其中缺失突变可以发生于HPRT基因的每个外显子 ,且诱发突变中大多数是多个外显子连锁缺失 (88 1% )。结论 ENU诱发HPRT基因突变图谱与自发突变完全不同 ,易诱发较大遗传结构改变 。

Objective To explore the molecular spectra and mechanism of human hypoxanthine guanine phosphoribosyl transferase (HPRT) gene mutation induced by ethyluitrosourea (ENU) Methods Single cell cloning culture, two way screening, multiple PCR amplification and electrophoresis technique were used Results With dose of ENU increasing, cell plating efficiency reduced (in the group with 100-200 μg/ml doses, P <0 01) and mutation frequency increased (in the group with 12 5-200 0 μg/ml doses, P <0 05) significantly There was no all exons deletion in spontaneous mutations, and only 7 7% of them were detected as single exon deletion But, deletion was found in 79 7% of ENU induced mutations (62 5%-89 4%, P <0 01), and deletion mutations in all nine exons of HPRT gene Most of ENU induced mutations were chain deletion with multiple exons (88 1%) Conclusions The spectra in spontaneous mutations differed completely from ENU induced ones ENU was liable to cause bigger changes in genetic structure, which suggested a stronger ENU′s mutagenesis