中国组织工程研究

中国组织工程研究 ›› 2010, Vol. 14 ›› Issue (24): 4430-4434.doi: 10.3969/j.issn.1673-8225.2010.24.014

• 组织构建实验造模 experimental modeling in tissue construction • 上一篇    下一篇

四种石蜡包埋组织DNA提取方法的比较

袁亚婷1，江岳鑫1，尹小文2，江兴堂2   

  1. 1厦门大学医学院，福建省厦门市   361004；2厦门大学附属中山医院呼吸内科，福建省厦门市  361004
  • 出版日期:2010-06-11 发布日期:2010-06-11
  • 通讯作者: 江兴堂，教授，厦门大学附属中山医院呼吸内科，福建省厦门市 361004 xmzsjxt@163.com
  • 作者简介:袁亚婷★，女，1983年生，山东省成武县人，汉族，厦门大学在读硕士，主要从事非小细胞肺癌表皮生长因子受体基因突变的研究。 yyting_123@163.com

Comparison of four methods for DNA extraction from paraffin-embedded tissues

Yuan Ya-ting1, Jiang Yue-xin1, Yin Xiao-wen2, Jiang Xing-tang2   

  1. 1 Medical College of Xiamen University, Xiamen  361004, Fujian Province, China; 2 Department of Respiratory Medicine, Zhongshan Hospital, Xiamen University, Xiamen  361004, Fujian Province, China
  • Online:2010-06-11 Published:2010-06-11
  • Contact: Jiang Xing-tang, Professor, Department of Respiratory Medicine, Zhongshan Hospital, Xiamen University, Xiamen 361004, Fujian Province, China xmzsjxt@163.com
  • About author:Yuan Ya-ting★, Studying for master’s degree, Medical College of Xiamen University, Xiamen 361004, Fujian Province, China yyting_123@163.com

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摘要：

背景：由于在甲醛固定石蜡包埋组织的制作及保存过程中对DNA造成的损害，影响了后续的聚合酶链反应等研究。选择一种简便有效且经济实用的石蜡包埋组织DNA提取方法成为研究者们关注和亟待解决的问题。
目的：比较甲醛固定石蜡包埋组织中4种提取DNA的方法对DNA质量的影响，探讨一种操作简便、污染少、经济实用的石蜡包埋组织中提取DNA的方法。
方法：取手术切除的普通甲醛固定石蜡包埋的非小细胞肺癌组织标本20例，分别以二甲苯脱蜡-酚氯仿法、改良TES水浴脱蜡-酚氯仿法、试剂盒法和改良TES水浴脱蜡-试剂盒抽提DNA法提取其DNA，然后进行电泳分析、紫外分光光度计测定A260/A280比值及PCR扩增。
结果与结论：改良TES水浴脱蜡-酚氯仿法和改良TES水浴脱蜡-试剂盒抽提DNA法可获得较大的DNA片段，且两种方法与试剂盒法所提取DNA的A260/A280值相比较，均有显著性意义(P < 0.05)，4种方法的提取效率差异无显著性意义(P > 0.1)，以改良TES水浴脱蜡-酚氯仿法所得DNA为模板，扩增的目的条带亮度与阳性对照相当。结果证实改良TES水浴脱蜡-酚氯仿法简便有效，所用试剂价格低廉，是一种经济实用的石蜡包埋组织DNA提取方法。

关键词: 甲醛, 石蜡包埋组织, DNA提取

Abstract:

BACKGROUND: Because of the damage of formalin-fixed paraffin-embedded tissue (FFPET) to DNA during the production and preservation, it is difficult to extract high quality and sufficient DNA for the following researches such as polymerase chain reaction (PCR). Therefore, it is essential and urgent to investigate an effective and economic method for extraction of DNA from FFPET.
OBJECTIVE: To investigate the influence of four DNA extraction methods for FFPET on DNA quality, and to identify the optimal method for DNA extraction from FFPET.
METHODS: Twenty formalin-fixed, paraffin-embedded non-small cell lung cancer (NSCLC) tissue specimens were selected and divided into four groups according to the method for DNA extraction. Four methods included xylene dewaxing and phenol-chloroform method, improved TES dewaxing with water-bath and phenol-chloroform method, genomic DNA extraction kit, improved TES dewaxing with water-bath and genomic DNA extraction kit. The quality of obtained DNA was analyzed by electrophoresis and PCR amplification.
RESULTS AND CONCLUSION: For paraffin-embedded tissue specimens, better DNA fragments could be obtained by using the improved TES dewaxing with water-bath and phenol-chloroform method and improved TES dewaxing with water-bath and genomic DNA extraction kit. The A260/A280 ratio of DNA obtained by using improved TES dewaxing with water-bath and phenol-chloroform method and improved TES dewaxing with water-bath and genomic DNA extraction kit were statistically different from genomic DNA extraction kit (P < 0.01). There was no significant difference in efficiency among four mehtods (P > 0.1). The aim trips were as bright as the positive control when using the DNA extraction by improved TES dewaxing with water-bath and phenol-chloroform method as the template. The results demonstrated that improved TES dewaxing with water-bath and phenol-chloroform method is an optimal method for extraction of DNA from FFPET.

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