Kraytsberg Yevgenya, Guo Xinhong, Tao Saisai, Kuznetsov Alexandra, MacLean Catherine, Ehrlich Daniel, Feldman Evan, Dombrovsky Igor, Yang Deye, Cloutier Gregory J, Castaneda-Sceppa Carmen, Khrapko Konstantin
Beth Israel Deaconess Medical Center, Boston, MA, USA.
College of Biology, Hunan University, Changsha, People's Republic of China.
Methods Mol Biol. 2016;1351:33-46. doi: 10.1007/978-1-4939-3040-1_4.
Quantification of deletions in mtDNA is a long-standing problem in mutational analysis. We describe here an approach that combines the power of single-molecule PCR of the entire mitochondrial genome with the enrichment of the deletions by restriction digestion. This approach is indispensable if information about wide range of deletion types in a sample is critical, such as in studies concerning distribution of deletion breakpoints (as opposed to approaches where fraction of a single deletion or a limited set of deletions is used as a proxy for total deletion load). Because deletions in a sample are quantified almost exhaustively, the other important application of this approach involves studies where only small amounts of tissue, such as biopsies, are available.
线粒体DNA缺失的定量分析是突变分析中一个长期存在的问题。我们在此描述一种方法，该方法将对整个线粒体基因组进行单分子PCR的能力与通过限制性消化富集缺失相结合。如果样本中广泛的缺失类型信息至关重要，例如在有关缺失断点分布的研究中（与将单一缺失或有限一组缺失的比例用作总缺失负荷替代指标的方法相反），那么这种方法是必不可少的。由于样本中的缺失几乎能被详尽地定量，该方法的另一个重要应用涉及仅能获得少量组织（如活检组织）的研究。
