坏变细胞的DNA对HpaⅡ限制性片段分析法测量DNA甲基化水平的干扰作用  

INTERFERING EFFECT OF DNA FROM DEGENERATING CELLS ON THE MEASUREMENT OF DNA METHYLATION ON LEVEL BY HPAⅡ RESTRICTION FRAGMENT ANALYSIS METHOD

作　　者：潘孔寒[1] 施正政[1] 余应年[1] 陈星若[1] 

机构地区：[1]浙江医科大学病理生理学教研室

出　　处：《浙江医科大学学报》1992年第2期49-53,共5页

基　　金：国家自然科学基金

摘　　要：用~3H-TdR在不同时期标记细胞DNA,以区别在HpaⅡ限制性片段放射活性分析中的DNA系来自增殖中的或坏变中的细胞.实验证明MNNG处理后存活的细胞中并无DNA甲基化水平的变化.而经MNNG或Hanks液处理的脱落细胞DNA,在HpaⅡ水解前后其DNA片段的L_w皆明显降低,并可干扰对新复制DNA的HpaⅡ识别位点的分析.因此作者认为,在缺乏严格设计以除外上述因素的体系中,作出关于细胞DNA甲基化状态变化的结论是不可信的.This paper reports that the Lvalue (kb) of HpaⅡ digests of cellular DNA labelled with ~3H-TdR before harvest was not changed in the surviving FL cells after MNNG treatment. While in FL cells which had been prelabelled with ~3H-TdR and then treated with MNNG or Hanks so- lution, the L?s of DNA from detached cells were significantly lower than that of the control. After HpaⅡ digestion, the L?s of restriction fragments were remarkably lower than that of the controls. The L?s of HpaⅡ digests of the mixed DNAs of the detached cells were lower than that of the control, after Hanks solution treatment with the control cells and those of the mixed DNAs of surviving cells with the detached cells after MNNG exposure were also lower than that of the controls. Thus, it has been clearly demonstrated that the presence of degenerating cells whose DNA has been broken by autolysis and/or chemical mutagen exposure could affect the result of DNA methylation when specific endonuclease cleavage site analysis is adopted. So the conclusion about cellular DNA hypomethylation could be reached only through experimental design that could restrict the object to be analyzed only in newly-replicated cellular DNA and rule out the unstable and unheritable DNA hypomethylation.

关 键 词：细胞 脱氢核糖核酸 甲基化 致癌物 

分 类 号：R730.21[医药卫生—肿瘤]