Simultaneous quantification of DNA damage and mitochondrial copy number by long-run DNA-damage quantification (LORD-Q).
Simultaneous quantification of DNA damage and mitochondrial copy number by long-run DNA-damage quantification (LORD-Q).
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DOI：
10.18632/oncotarget.20112
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发表时间：
2017-12-22
期刊：
影响因子：
--
通讯作者：
Schulze-Osthoff K
中科院分区：
文献类型：
--
作者：
Dannenmann B;Lehle S;Lorscheid S;Huber SM;Essmann F;Schulze-Osthoff K
DNA损伤和线粒体DNA含量的变化与衰老和癌症的发展有关。为了防止基因组不稳定和肿瘤发生，细胞必须保持其核和线粒体DNA的完整性。然而，DNA损伤保护和基因组稳定性研究的进展也依赖于能够以准确的高通量方式可靠地量化线粒体DNA拷贝数变化和DNA损伤的技术的可用性。不幸的是，目前还没有建立这样的方法。在这里，我们描述了用于DNA损伤定量的高灵敏度长期实时PCR技术（LORD-Q）及其在培养细胞和组织样本中同时测量DNA损伤率和线粒体DNA拷贝数的适用性。使用LORD-Q多重试验，我们举例说明线粒体DNA含量不直接影响DNA损伤易感性，但影响某些抗癌药物的疗效。因此，LORD-Q提供了一种快速、精确的方法来评估DNA损伤、DNA修复和mtDNA复制，以及它们在各种病理环境中的作用。
DNA damage and changes in the mitochondrial DNA content have been implicated in ageing and cancer development. To prevent genomic instability and tumorigenesis, cells must maintain the integrity of their nuclear and mitochondrial DNA. Advances in the research of DNA damage protection and genomic stability, however, also depend on the availability of techniques that can reliably quantify alterations of mitochondrial DNA copy numbers and DNA lesions in an accurate high-throughput manner. Unfortunately, no such method has been established yet. Here, we describe the high-sensitivity long-run real-time PCR technique for DNA-damage quantification (LORD-Q) and its suitability to simultaneously measure DNA damage rates and mitochondrial DNA copy numbers in cultured cells and tissue samples. Using the LORD-Q multiplex assay, we exemplarily show that the mitochondrial DNA content does not directly affect DNA damage susceptibility, but influences the efficacy of certain anticancer drugs. Hence, LORD-Q provides a fast and precise method to assess DNA lesions, DNA repair and mtDNA replication as well as their role in a variety of pathological settings.
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