四种微量石蜡组织DNA提取方法的比较_临床医学论文.docVIP

四种微量石蜡组织DNA提取方法的比较_临床医学论文 四种微量石蜡组织DNA提取方法的比较_临床医学论文 作者：黄幼生，宋伟伟，邓晓佳，罗志飞，解娜 【摘要】 目的：探讨从微量福尔马林固定石蜡包埋组织（formalin fixation and paraffin embedded tissues，FFPET)中提取DNA的简单而高效的方法。方法：采用Chelex提取法、酚氯仿抽提法、单纯消化法、水煮法4种方法提取组织DNA；应用聚合酶链反应扩增100～500bp长度DNA片段，然后进行电泳分析，1年后重复PCR电泳分析。结果：小于200bp长度DNA片段扩增，Chelex提取法、单纯消化法阳性率分别为88.8%、78.8%，明显优于酚氯仿抽提法（15.0%）（0.01）及水煮法（21.3%）（0.01）。随着扩增长度的增加，PCR扩增效率逐渐降低，Chelex提取法PCR反应总阳性率为82.5%，单纯消化法为66.5%，水煮法为13.4%，酚氯仿抽提法为13.4%。1年后重复PCR总阳性率分别为80.1%、31.7%、2.5%、9.2%。结论：Chelex提取法方便简单，适用于微量石蜡组织DNA扩增分析；单纯消化法及水煮法在一定条件下适用于微量石蜡组织短片段DNA的扩增。 【关键词】 石蜡组织;DNA提取;Chelex提取法［ABSTRACT］ Objective: To explore the simple and effective method for DNA extraction from microamount formalin fixation and paraffin embedded tissues (FFPET). Methods: The DNA was extracted by four different methods: Chelexysis was repeated 1 year later. Results: For amplification of DNA less than 200 bp, the positive rates of Chelex100 group and simple digestion group were 88.8% and 78.8%, respectively, which were significantly effective than phenolchloroform purification group (15.0%) (0.01) and water cooking group (21.3%) (0.01). The amplication efficacy decreased as the length increase. The total positive rates of PCR were 82.5% in Chelex100 group, 66.5% in simple digestion group, 13.4% both in water cooking group and phenolchlorform group. And the positive rates in repeated assessment were 80.1%, 31.7%, 2.5% and 9.2%. Conclusions: Chelex100 extration method is easy and applicable in amplification analysis of DNA extracted from mircoamount paraffin embedded tissues; while simple digestion method and water cooking method are suitable to analysis of short length DNA fragment extracted from mircoamount paraffin embedded tissues. ［KEY WORDS］ Paraffin embedded tissue; DNA extraction; Chelex 医院病理科存在大量各种疾病甲醛固定石蜡包埋组织（formalin fixed and paraffin embedded tissues， FFPET），为分子病理研究提供了大量的信息来源。然而福尔马林固定后的蜡块包埋组织DNA降解严重［1］，档存FFPET组织切取量有限，传统酚氯仿提取DNA步骤繁杂，提取过程损失严重等都制约了石蜡组织在分子病理学中的研究应用［2］。如何高效、高质量提取微量石蜡组织DNA对利用石蜡组织进行