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昆明山海棠诱导人白血病细胞HPRT基因突变的研究 |
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刘胜学 曹佳 安辉 周紫垣
摘 要：为研究昆明山海棠(THH)的遗传毒性和药理作用，采用单细胞克隆培养,双向筛选计数，多重PCR扩增与电泳分析，研究了THH诱导HL－60细胞HPRT基因突变率及分子突变谱。发现随着染毒剂量的增加，细胞接种存活率逐渐下降，突变频率明显升高；THH诱发突变主要由缺失和点突变两部分组成(46.6％和53.4％)，而自发突变几乎全是点突变(92.3％)；HPRT基因突变位点在各个外显子的分布较集中于基因的3′末端，且外显子1缺失只出现于全基因缺失中，外显子7/8与9多表现为连锁缺失(71.4％)。结果提示，THH具有明确的诱导HPRT基因突变的作用，且诱发突变与自发突变的分子图谱不一样，这可能与其作用机制有关。上述发现有助于阐明THH遗传毒性作用机理。
关键词：HPRT基因;昆明山海棠;致突变作用;HL－60细胞;多重PCR
中图分类号：R733.72 文献标识码：A
文章编号：0253－9772(2000)05－0305－04
Molecular Analysis of THH-Induced Mutations at the HPRT
Locus in Human Promyelocytic Leukemia Cells by
Multiplex Polymerase Chain Reaction
LIU Sheng-xue，CAO Jia，AN Hui，ZHOU Zi-yuan
(Department of Molecular Toxicololgy,Third Military Medical University,Chongqing 400038,China)
Abstract:The genotoxicity and pharmacologic activity of a Chinese medicinal herb,Tripterygium hypoglaucum(Lévl) Hutch (THH),was investigated by methods of single cell clone culture,two-way screening count,multiplex PCR amplification and electrophoresis technique.THH showed clear cytotoxicity and mutagenesis in human promyelocytic leukemia (HL-60) cells.When doses were increased,cell plating efficiency reduced and mutation frequency increased.The analysis showed that the spectra of spontaneous and THHinduced mutants were different.46.6％ of THHinduced genetic changes were deletions,whereas the majority of spontaneous mutants(92.3％)exhibited point mutations.Mapping of all intragenic deletion breakpoints showed a random distribution of breakpoints in 9 exons,but toward the 3′ end of the HPRT gene.Exon 1 deletion only appeared in total gene deletion,and exon 7/8 and 9 deletion often showed chain deletion(71.4％).
Key words:HPRT gene;Tripterygium hypoglaucum Hutch;mutagenesis;HL－60 cells;multiplex PCR
昆明山海棠(Tripterygium hypoglaucum (Lévl) Hutch,THH)为卫茅科雷公藤属植物，主要化学成分包括生物碱，萜类及色素剂，在中医学中常用于治疗风湿性关节炎，红斑狼疮等自身免疫性疾病，由于发现THH具有可逆性抗生育及抗肿瘤[1] [2] 下一页
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