Institute for Organic Synthesis and Photoreactivity, National Research Council, 40129 Bologna, Italy; Center of Advanced Technologies, Adam Mickiewicz University, 61-712 Poznań, Poland.
Institute of Nanoscience and Nanotechnology, National Center for Scientific Research Demokritos, 15310 Athens, Greece.
STAR Protoc. 2024 Sep 20;5(3):103191. doi: 10.1016/j.xpro.2024.103191. Epub 2024 Aug 14.
Most DNA damages induced through oxidative metabolism are single lesions which can accumulate in tissues. Here, we present a protocol for the simultaneous quantification of oxidative purine lesions (cPu and 8-oxo-Pu) in DNA. We describe steps for enzymatic digestion of DNA and sample pre-purification, followed by quantification through liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. We optimized this protocol in commercially available calf thymus DNA and used genomic and mitochondrial DNA extracted from cell cultures and animal and human tissues.
大多数通过氧化代谢诱导的 DNA 损伤是单损伤，可以在组织中积累。在这里，我们提出了一种同时定量 DNA 中氧化嘌呤损伤（cPu 和 8-oxo-Pu）的方案。我们描述了酶消化 DNA 和样品预纯化的步骤，然后通过液相色谱-串联质谱（LC-MS/MS）分析进行定量。我们在市售的小牛胸腺 DNA 中优化了该方案，并使用细胞培养物以及动物和人体组织中提取的基因组和线粒体 DNA 进行了验证。
