Shaw Andrew M, Gammage Payam A
CRUK Beatson Institute, Glasgow, UK.
Institute of Cancer Sciences, University of Glasgow, Glasgow, UK.
Methods Mol Biol. 2023;2615:31-40. doi: 10.1007/978-1-0716-2922-2_3.
Direct analysis of mtDNA using PCR-free methods is limited by the presence of persistent, contaminating nucleic acids originating from the nuclear genome, even following stringent mitochondrial isolations. Here we describe a method developed in our laboratory that couples existing, commercially available mtDNA isolation protocols with exonuclease treatment and size exclusion chromatography (DIFSEC). This protocol produces highly enriched mtDNA extracts from small-scale cell culture, with near-undetectable nuclear DNA contamination.
即使经过严格的线粒体分离，使用无PCR方法直接分析线粒体DNA（mtDNA）仍受到源自核基因组的持久性污染核酸的限制。在此，我们描述了一种在我们实验室开发的方法，该方法将现有的市售mtDNA分离方案与核酸外切酶处理和尺寸排阻色谱法（DIFSEC）相结合。该方案从小规模细胞培养中产生高度富集的mtDNA提取物，核DNA污染几乎检测不到。
