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Retrieving high-quality genomic DNA from formalin-fixed paraffin-embedded tissues for multiple molecular analyses
Preparative Biochemistry & Biotechnology ( IF 2.0 ) Pub Date : 2021-05-28 , DOI: 10.1080/10826068.2021.1923030
Ha Thi Nguyen 1, 2 , Vinay Bharadwaj Tatipamula 1, 2 , Duy Ngoc Do 3 , Thien Chi Huynh 4 , Mai Kim Dang 1
Affiliation  

Abstract

Formalin-fixed paraffin-embedded (FFPE) tissues represent the biggest source of archival materials for molecular biology research and pathology investigations. Nevertheless, fixation by formalin may cause denaturation and modification of macromolecules constraining DNA quality and its downstream applications. In this study, we developed a fast, simple, and cost-effective phenol/chloroform-based protocol for the extraction of high-quality DNA from 101 FFPE colorectal cancer tissue blocks that can be used in multiple molecular studies. DNA samples extracted using this phenol/chloroform protocol and the QIAamp DNA FFPE Tissue kit were evaluated for the quantity, quality, and amplificability. Spectrophotometer analyses revealed significantly higher quality and quantity of DNA samples obtained with the phenol/chloroform protocol as compared to those of the QIAamp DNA FFPE Tissue kit. In addition, the amplificability of these samples as assessed by conventional and multiplex polymerase chain reaction (PCR), followed by sequencing and fragment analyses presented an absolute success rate. Additionally, it is able to amplify a DNA fragment of 725 base-pairs at an adequate amount for downstream applications. This fast, simple, and cost-effective method may facilitate the molecular analyses of a large number of FFPE specimens that best suits the needs of the overall study design in terms of the quality and quantity of the extracted DNA.



中文翻译:

从福尔马林固定的石蜡包埋组织中提取高质量基因组 DNA 以进行多种分子分析

摘要

福尔马林固定石蜡包埋 (FFPE) 组织是分子生物学研究和病理学调查档案材料的最大来源。然而,福尔马林固定可能会导致限制 DNA 质量及其下游应用的大分子的变性和修饰。在本研究中,我们开发了一种快速、简单且经济高效的苯酚/氯仿方案,用于从 101 个 FFPE 结直肠癌组织块中提取高质量 DNA,可用于多种分子研究。对使用此苯酚/氯仿方案和 QIAamp DNA FFPE Tissue 试剂盒提取的 DNA 样品的数量、质量和扩增能力进行了评估。分光光度计分析显示,与 QIAamp DNA FFPE Tissue 试剂盒相比,使用苯酚/氯仿方案获得的 DNA 样品的质量和数量明显更高。此外,通过常规和多重聚合酶链反应 (PCR) 评估的这些样品的扩增能力,然后进行测序和片段分析,显示出绝对成功率。此外,它能够以足够的量扩增 725 个碱基对的 DNA 片段,以供下游应用使用。这种快速、简单且具有成本效益的方法可以促进大量 FFPE 样本的分子分析,在提取 DNA 的质量和数量方面最适合整体研究设计的需求。通过常规和多重聚合酶链反应 (PCR) 评估的这些样品的扩增能力,然后进行测序和片段分析,显示出绝对成功率。此外,它能够以足够的量扩增 725 个碱基对的 DNA 片段,以供下游应用使用。这种快速、简单且具有成本效益的方法可以促进大量 FFPE 样本的分子分析,在提取 DNA 的质量和数量方面最适合整体研究设计的需求。通过常规和多重聚合酶链反应 (PCR) 评估的这些样品的扩增能力,然后进行测序和片段分析,显示出绝对成功率。此外,它能够以足够的量扩增 725 个碱基对的 DNA 片段,以供下游应用使用。这种快速、简单且具有成本效益的方法可以促进大量 FFPE 样本的分子分析,在提取 DNA 的质量和数量方面最适合整体研究设计的需求。

更新日期:2021-05-28
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