Abicht Angela, Scharf Florentine, Kleinle Stephanie, Schön Ulrike, Holinski-Feder Elke, Horvath Rita, Benet-Pagès Anna, Diebold Isabel
Medical Genetic Center Munich, Munich, Germany.
Department of Neurology, Friedrich-Baur-Institute, Klinikum der Ludwig-Maximilians-Universität München, Munich, Germany.
Mol Genet Genomic Med. 2018 Nov;6(6):1188-1198. doi: 10.1002/mgg3.500. Epub 2018 Nov 8.
The diagnosis of mitochondrial disorders is challenging because of the clinical variability and genetic heterogeneity of these conditions. Next-Generation Sequencing (NGS) technology offers a robust high-throughput platform for nuclear and mitochondrial DNA (mtDNA) analyses.
We developed a custom Agilent SureSelect Mitochondrial and Nuclear Disease Panel (Mito-aND-Panel) capture kit that allows parallel enrichment for subsequent NGS-based sequence analysis of nuclear mitochondrial disease-related genes and the complete mtDNA genome. Sequencing of enriched mtDNA simultaneously with nuclear genes was compared with the separated sequencing of the mitochondrial genome and whole exome sequencing (WES).
The Mito-aND-Panel permits accurate detection of low-level mtDNA heteroplasmy due to a very high sequencing depth compared to standard diagnostic procedures using Sanger sequencing/SNaPshot and WES which is crucial to identify maternally inherited mitochondrial disorders.
We established a NGS-based method with combined sequencing of the complete mtDNA and nuclear genes which enables a more sensitive heteroplasmy detection of mtDNA mutations compared to traditional methods. Because the method promotes the analysis of mtDNA variants in large cohorts, it is cost-effective and simple to setup, we anticipate this is a highly relevant method for sequence-based genetic diagnosis in clinical diagnostic applications.
线粒体疾病的诊断具有挑战性，因为这些病症存在临床变异性和遗传异质性。下一代测序（NGS）技术为核DNA和线粒体DNA（mtDNA）分析提供了一个强大的高通量平台。
我们开发了一种定制的安捷伦SureSelect线粒体和核疾病检测试剂盒（Mito-aND-Panel）捕获试剂盒，该试剂盒允许对核线粒体疾病相关基因和完整的mtDNA基因组进行平行富集，以便后续基于NGS的序列分析。将富集的mtDNA与核基因同时测序与线粒体基因组单独测序和全外显子组测序（WES）进行比较。
与使用桑格测序/SNaPshot和WES的标准诊断程序相比，Mito-aND-Panel由于测序深度非常高，能够准确检测低水平的mtDNA异质性，这对于识别母系遗传的线粒体疾病至关重要。
我们建立了一种基于NGS的方法，对完整的mtDNA和核基因进行联合测序，与传统方法相比，该方法能够更灵敏地检测mtDNA突变。由于该方法促进了对大量队列中mtDNA变异的分析，具有成本效益且设置简单，我们预计这是一种在临床诊断应用中基于序列的基因诊断的高度相关方法。
