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存档福尔马林固定石蜡包埋组织中 DNA 提取和纯化方案的效率评估。

Efficiency evaluation of a DNA extraction and purification protocol on archival formalin-fixed and paraffin-embedded tissue.

机构信息

Institute of Legal Medicine, EA4438, University of Strasbourg, Strasbourg, France.

出版信息

Forensic Sci Int. 2010 Jan 30;194(1-3):e25-8. doi: 10.1016/j.forsciint.2009.09.004. Epub 2009 Sep 24.

Abstract

Formalin-fixed and paraffin-embedded tissue (FF-PET) is an invaluable resource for retrospective molecular genetic studies, but the extraction of high-quality genomic DNA from FF-PET is still a problematic issue. Despite the range of DNA extraction methods currently in use, the association of phenol-chloroform extraction and silica-based purification protocols, reported in ancient DNA studies on archaeological bones, has, to our knowledge, not been used for DNA extraction from FF-PET yet. The present study compared the efficiency of three DNA extraction and purification protocols from two different FF-PET substrates, heart and liver, by using quantitative PCR and multiplex amplification. We showed that the method, using phenol-chloroform and the QIAamp DNA mini Kit (Qiagen), was the most effective DNA extraction and purification method and that the DNA quantity extracted from liver is statistically more important than that extracted from heart. Autosomal STR typing by multiplex amplifications gave partial allelic profiles with only small size products (less than 300 bases) amplified, suggesting that DNA extracted from FF-PET was degraded. In conclusion, the protocol presented here, previously described in studies on ancient bones, should find application in different molecular studies involving FF-PET material.

摘要

福尔马林固定石蜡包埋组织(FF-PET)是回顾性分子遗传学研究的宝贵资源,但从 FF-PET 中提取高质量的基因组 DNA 仍然是一个有问题的问题。尽管目前有多种 DNA 提取方法,但在对考古骨骼的古代 DNA 研究中报告的酚-氯仿提取和基于硅的纯化方案的关联,据我们所知,尚未用于从 FF-PET 中提取 DNA。本研究通过定量 PCR 和多重扩增比较了从两种不同的 FF-PET 基质(心脏和肝脏)中使用三种 DNA 提取和纯化方案的效率。我们表明,使用酚-氯仿和 QIAamp DNA mini 试剂盒(Qiagen)的方法是最有效的 DNA 提取和纯化方法,并且从肝脏提取的 DNA 数量在统计学上比从心脏提取的 DNA 更重要。通过多重扩增进行的常染色体 STR 分型仅获得了小片段产物(小于 300 个碱基)扩增的部分等位基因谱,表明从 FF-PET 提取的 DNA 已降解。总之,这里提出的方案以前在对古代骨骼的研究中已有描述,应该可以应用于涉及 FF-PET 材料的不同分子研究。

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