石蜡包埋淋巴瘤组织线粒体DNA的提取及PCR扩增

    Extraction and PCR amplification of mitochondrial DNA from paraffin-embedded lymphoma tissue

    • 摘要: 目的:探讨从石蜡包埋组织标本获取线粒体DNA (mtDNA)并进行PCR扩增的有效方法。方法:选取27例石蜡包埋的淋巴瘤组织,分别使用二甲苯/乙醇和0.5% Tween-20进行脱蜡,改进裂解缓冲液的成分及浓度,酚/氯仿抽提以获取mtDNA并进行电泳和PCR分析。结果:0.5% Tween-20法获取mtDNA纯度及浓度含量明显高于二甲苯/乙醇法(P<0.01);二甲苯/乙醇法和0.5% Tween-20法获取的mtDNA PCR扩增成功率分别为11.1%和40.7%,两者差异有统计学意义(P<0.05)。结论:二甲苯/乙醇和0.5% Tween-20两种方法都可以扩增出mtDNA的目的片段,但0.5% Tween-20法操作简便,PCR扩增成功率高,不用接触二甲苯等有毒化学试剂,是一种较为理想的mtDNA提纯方法。

       

      Abstract: Objective: To explore an effective method for extraction and PCR amplification of mitochondrial DNA from paraffin-embedded lymphoma tissue.Methods: Twenty seven paraffin-embedded lymphoma tissue specimens were selected.The two methods:Xylene/ethanol method and 0.5% Tween-20 method were used for deparaffinisation.The component and concentration of lysis buffer were modified,and mitochondrial DNA was extracted by phend and chloroform.The obtained mitochondrial DNA was analyzed by PCR amplification.Results: The DNA purity and concentration which extracted by 0.5% Tween-20 deparaffinisation method was obviously higher than that by Xylene/ethanol deparaffinisation method.The successrate of PCR amplification was 11.1% by Xylene/ethanol method and 40.7% by 0.5% Tween-20 method,respectively.There was significant statistical difference (P<0.05).Conclusions: Both the Xylene/ethanol deparaffinisation method and 0.5% Tween-20 deparaffinisation method can extract mitochondrial DNA.But the 0.5% Tween-20 method is simple,the success rate is high,and can reduce the contact chance from poisonous chemical reagents such as dimethylbenzene.So the 0.5% Tween-20 deparaffinisation method is an ideal method for extraction of mitochondrial DNA from paraffin-embedded tissues.

       

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