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利用 phi29 DNA 聚合酶进行 DNA 扩增可验证从口腔黏膜样本中进行的基因多态性分析。

DNA amplification using phi29 DNA polymerase validates gene polymorphism analysis from buccal mucosa samples.

机构信息

Department of Oral Reconstruction and Rehabilitation, Kyushu Dental College, Graduate School, Japan.

出版信息

J Prosthodont Res. 2011 Jul;55(3):165-70. doi: 10.1016/j.jpor.2010.12.001. Epub 2011 Feb 5.

Abstract

Venous blood is currently the most common source of DNA for gene polymorphism screening; however, blood sampling is invasive and difficult to perform in general dental treatment. Buccal mucosa samples provide an alternative source of DNA, but it is frequently difficult to effectively amplify the DNA owing to the small amounts of sample material obtained. This study was performed to establish a method for performing total genomic DNA amplification from buccal mucosa samples using phi29 DNA polymerase. Total genomic DNA was isolated from buccal mucosa samples obtained from healthy subjects and was amplified using phi29 DNA polymerase. To determine the suitability of the extracted DNA for genotyping, polymerase chain reaction and restriction fragment length polymorphism analyses were performed for the IL-1 gene polymorphism. Genotyping of the IL-1 polymorphism was successful using the amplified DNA from a buccal mucosa, but genotyping was unsuccessful using the unamplified control because of low DNA purity. The method of extracting DNA from a buccal mucosa is painless, simple, minimally invasive, and rapid. Genomic DNA from a buccal mucosa can be amplified by phi29 DNA polymerase in sufficient quantity and quality to conduct gene polymorphism analyses.

摘要

静脉血是目前用于基因多态性筛选的最常见的 DNA 来源;然而,一般的牙科治疗中,采血是具有侵入性的,且难以进行。颊黏膜样本提供了 DNA 的另一种来源,但由于获得的样本材料量小,通常难以有效地扩增 DNA。本研究旨在建立一种使用 phi29 DNA 聚合酶从颊黏膜样本中进行全基因组 DNA 扩增的方法。从健康受试者的颊黏膜样本中分离出全基因组 DNA,并使用 phi29 DNA 聚合酶进行扩增。为了确定提取的 DNA 用于基因分型的适用性,对 IL-1 基因多态性进行了聚合酶链反应和限制性片段长度多态性分析。使用从颊黏膜扩增的 DNA 成功进行了 IL-1 多态性的基因分型,但未使用未扩增的对照进行基因分型,因为 DNA 纯度低。从颊黏膜提取 DNA 的方法是无痛、简单、微创且快速的。phi29 DNA 聚合酶可足够数量和质量地扩增来自颊黏膜的基因组 DNA,以进行基因多态性分析。

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