Claudia P. Gonzalez-Hunt, John P. Rooney, Ian T. Ryde, Charumathi Anbalagan, Rashmi Joglekar, Joel N. Meyer
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引用次数: 72
Abstract
Because of the role that DNA damage and depletion play in human disease, it is important to develop and improve tools to assess these endpoints. This unit describes PCR-based methods to measure nuclear and mitochondrial DNA damage and copy number. Long amplicon quantitative polymerase chain reaction (LA-QPCR) is used to detect DNA damage by measuring the number of polymerase-inhibiting lesions present based on the amount of PCR amplification; real-time PCR (RT-PCR) is used to calculate genome content. In this unit, we provide step-by-step instructions to perform these assays in Homo sapiens, Mus musculus, Rattus norvegicus, Caenorhabditis elegans, Drosophila melanogaster, Danio rerio, Oryzias latipes, Fundulus grandis, and Fundulus heteroclitus, and discuss the advantages and disadvantages of these assays. © 2016 by John Wiley & Sons, Inc.
基于pcr的线粒体DNA拷贝数、线粒体DNA损伤和核DNA损伤分析
由于DNA损伤和耗竭在人类疾病中发挥的作用,开发和改进评估这些终点的工具非常重要。本单元描述了基于pcr的方法来测量核和线粒体DNA损伤和拷贝数。长扩增子定量聚合酶链反应(LA-QPCR)用于检测DNA损伤,基于PCR扩增量测量存在的聚合酶抑制病变的数量;实时荧光定量PCR (RT-PCR)用于计算基因组含量。在本单元中,我们提供了一步一步的说明,在智人,小家鼠,褐家鼠,秀丽隐杆线虫,黑腹果蝇,猕猴,大眼底和异眼底进行这些检测,并讨论了这些检测的优点和缺点。©2016 by John Wiley &儿子,Inc。
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