Comparison of methods in the recovery of nucleic acids from archival formalin-fixed paraffin-embedded autopsy tissues
Comparison of methods in the recovery of nucleic acids from archival formalin-fixed paraffin-embedded autopsy tissues
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DOI：
10.1016/j.ab.2010.01.014
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发表时间：
2010-05-01
影响因子：
2.9
通讯作者：
Poinar, Hendrik N.
中科院分区：
文献类型：
--
作者：
Okello, John B. A.;Zurek, Jaymi;Poinar, Hendrik N.
在发展中国家，档案福尔马林固定石蜡包埋 (FFPE) 人体组织收藏的储存状态通常很差。随着生物分子技术的进步，这些非凡且几乎未开发的资源已成为回顾性流行病学研究的重要组成部分。为了在基因组研究中成功使用此类组织，科学家需要高核酸产量和纯度。尽管可用的 FFPE 组织试剂盒数量不断增加，但很少有研究分析其从存档的人体尸检样本中回收高质量核酸的适用性。在这里，我们提供了一项涉及 10 种主要提取方法的研究，用于从年龄从 3 岁到 13 岁的 FFPE 组织中分离总核酸。尽管所有 10 种方法都回收了可定量的 DNA，但只有 6 种方法回收了可定量的 RNA，差异很大，并且通常产生较低的 DNA 浓度。总体而言，我们定量地表明，TrimGen 的 WaxFree 方法和我们内部的苯酚-氯仿提取方法回收了最高产率的可扩增 DNA，并具有相当大的聚合酶链式反应 (PCR) 抑制作用，而 Ambion 的 RecoverAll 方法回收了最高的可扩增 RNA。 (C) 2010 Elsevier Inc. 保留所有权利。
Archival formalin-fixed paraffin-embedded (FFPE) human tissue collections are typically in poor states of storage across the developing world. With advances in biomolecular techniques, these extraordinary and virtually untapped resources have become an essential part of retrospective epidemiological studies. To successfully use such tissues in genomic studies, scientists require high nucleic acid yields and purity. In spite of the increasing number of FFPE tissue kits available, few studies have analyzed their applicability in recovering high-quality nucleic acids from archived human autopsy samples. Here we provide a study involving 10 major extraction methods used to isolate total nucleic acid from FFPE tissues ranging in age from 3 to 13 years. Although all 10 methods recovered quantifiable amounts of DNA, only 6 recovered quantifiable RNA, varying considerably and generally yielding lower DNA concentrations. Overall, we show quantitatively that TrimGen's WaxFree method and our in-house phenol-chloroform extraction method recovered the highest yields of amplifiable DNA, with considerable polymerase chain reaction (PCR) inhibition, whereas Ambion's RecoverAll method recovered the most amplifiable RNA. (C) 2010 Elsevier Inc. All rights reserved.