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一种从人类样本中提取基因组DNA用于PCR-RFLP分析的简单方法。

A simple method of genomic DNA extraction from human samples for PCR-RFLP analysis.

作者信息

Ghatak Souvik, Muthukumaran Rajendra Bose, Nachimuthu Senthil Kumar

机构信息

Departments of Biotechnology and.

出版信息

J Biomol Tech. 2013 Dec;24(4):224-31. doi: 10.7171/jbt.13-2404-001.

Abstract

Isolation of DNA from blood and buccal swabs in adequate quantities is an integral part of forensic research and analysis. The present study was performed to determine the quality and the quantity of DNA extracted from four commonly available samples and to estimate the time duration of the ensuing PCR amplification. Here, we demonstrate that hair and urine samples can also become an alternate source for reliably obtaining a small quantity of PCR-ready DNA. We developed a rapid, cost-effective, and noninvasive method of sample collection and simple DNA extraction from buccal swabs, urine, and hair using the phenol-chloroform method. Buccal samples were subjected to DNA extraction, immediately or after refrigeration (4-6°C) for 3 days. The purity and the concentration of the extracted DNA were determined spectrophotometerically, and the adequacy of DNA extracts for the PCR-based assay was assessed by amplifying a 1030-bp region of the mitochondrial D-loop. Although DNA from all the samples was suitable for PCR, the blood and hair samples provided a good quality DNA for restriction analysis of the PCR product compared with the buccal swab and urine samples. In the present study, hair samples proved to be a good source of genomic DNA for PCR-based methods. Hence, DNA of hair samples can also be used for the genomic disorder analysis in addition to the forensic analysis as a result of the ease of sample collection in a noninvasive manner, lower sample volume requirements, and good storage capability.

摘要

从血液和口腔拭子中提取足够量的DNA是法医研究与分析不可或缺的一部分。本研究旨在确定从四种常见样本中提取的DNA的质量和数量,并估算随后PCR扩增所需的时间。在此,我们证明头发和尿液样本也可成为可靠获取少量适用于PCR的DNA的替代来源。我们开发了一种快速、经济高效且非侵入性的样本采集方法,并使用酚 - 氯仿法从口腔拭子、尿液和头发中进行简单的DNA提取。口腔样本在提取DNA时,可立即进行,也可在4 - 6°C冷藏3天后进行。通过分光光度法测定提取的DNA的纯度和浓度,并通过扩增线粒体D - 环的1030 bp区域来评估DNA提取物对基于PCR的检测的适用性。尽管所有样本的DNA都适用于PCR,但与口腔拭子和尿液样本相比,血液和头发样本为PCR产物的限制性分析提供了高质量的DNA。在本研究中,头发样本被证明是基于PCR方法的良好基因组DNA来源。因此,由于头发样本易于以非侵入性方式采集、样本量要求较低且储存能力良好,其DNA除用于法医分析外,还可用于基因组疾病分析。

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Evaluation of three methods for effective extraction of DNA from human hair.三种从人发中有效提取DNA方法的评估。
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