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基于 qPCR 的方法定量检测端粒 DNA 中氧化鸟嘌呤和其他 FPG 敏感碱基损伤。

A qPCR-based assay to quantify oxidized guanine and other FPG-sensitive base lesions within telomeric DNA.

机构信息

Nutritional Genomics and DNA Damage Research Group, Food and Nutritional Sciences, Commonwealth Scientific and Industrial Research Organization, Adelaide, Australia. nathan.o'

出版信息

Biotechniques. 2011 Dec;51(6):403-11. doi: 10.2144/000113788.

Abstract

Telomere shortening is an important risk factor for cancer and accelerated aging. However, it is becoming evident that oxidatively damaged DNA within the telomere sequence may also cause telomere dysfunction. Here we describe a reliable, cost-effective quantitative PCR (qPCR)-based method to measure the amount of oxidized residues within telomeric DNA that are recognized and excised by formamidopyridine DNA glycosylase (FPG). We also report that in an in vitro model of oxidative stress oxidized base lesions measured using this method are more prevalent within telomeric sequences. Furthermore, this method is sufficiently sensitive to detect changes in oxidative stress induced by zinc deficiency and hydrogen peroxide within the physiological range.

摘要

端粒缩短是癌症和加速衰老的一个重要风险因素。然而,越来越明显的是,端粒序列中氧化损伤的 DNA 也可能导致端粒功能障碍。在这里,我们描述了一种可靠且经济有效的定量 PCR(qPCR)方法,用于测量被 formamidopyridine DNA glycosylase(FPG)识别和切除的端粒 DNA 中的氧化残基的量。我们还报告说,在体外氧化应激模型中,使用这种方法测量的氧化碱基损伤在端粒序列中更为普遍。此外,该方法足够灵敏,可以检测到生理范围内缺锌和过氧化氢诱导的氧化应激的变化。

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