Preventing the pollution of mitochondrial datasets with nuclear mitochondrial paralogs (numts)
Preventing the pollution of mitochondrial datasets with nuclear mitochondrial paralogs (numts)
复制标题
DOI：
10.1016/j.mito.2010.10.004
复制
发表时间：
2011-03-01
期刊：
影响因子：
4.4
通讯作者：
Douady, Christophe J.
中科院分区：
文献类型：
--
作者：
Calvignac, Sebastien;Konecny, Lara;Douady, Christophe J.
分子工具在生态学和进化中已经变得突出。分子生态学家和进化论者选择的一个目标是线粒体DNA（mtDNA），它的许多优点也说服了广泛的，务实的计划，如条形码倡议。当然，mtDNA也是研究线粒体疾病的人类遗传学家感兴趣的。然而，使用mtDNA的研究由于核假基因（numts）的无意共扩增或优先扩增而面临巨大风险。对假定的mtDNA序列进行后验分析可以帮助去除numts，但面临严重的限制（例如，最近易位的numts大多数时间不会被注意到）。先验采取的对策，即明确设计用于避免numt共扩增或优选扩增，是有吸引力的，但从未被适当评估。在这里，我们调查了四个这样的措施（mtDNA富集，cDNA扩增，远程扩增和PCR前稀释）的效率在一组常见的numt情况下，显示mtDNA富集是最差的表现，而使用PCR前稀释是一个简单的，但强大的方法，以防止污染的推定的mtDNA数据集与numts。因此，可以提出直接的建议，如果遵循，将大大增加任何mtDNA样序列的线粒体起源的信心。(C)2010 Elsevier B.V.和线粒体研究学会。All rights reserved.
Molecular tools have become prominent in ecology and evolution. A target of choice for molecular ecologists and evolutionists is mitochondrial DNA (mtDNA), whose many advantages have also convinced broad-scale, pragmatic programmes such as barcode initiatives. Of course, mtDNA is also of interest to human geneticists investigating mitochondrial diseases. Studies using mtDNA are however put at great risk by the inadvertent co-amplification or preferred amplification of nuclear pseudogenes (numts). A posteriori analysis of putative mtDNA sequences can help in removing numts but faces severe limitations (e.g. recently translocated numts will most of the time go unnoticed). Counter-measures taken a priori, i.e. explicitly designed for avoiding numt co-amplification or preferred amplification, are appealing but have never been properly assessed. Here we investigate the efficiency of four such measures (mtDNA enrichment, cDNA amplification, long-range amplification and pre-PCR dilution) on a common set of numt cases, showing that mtDNA enrichment is the worst performer while the use of pre-PCR dilution is a simple, yet robust method to prevent the pollution of putative mtDNA datasets with numts. Therefore, straightforward recommendations can be made that, if followed, will considerably increase the confidence in the mitochondrial origin of any mtDNA-like sequence. (C) 2010 Elsevier B.V. and Mitochondria Research Society. All rights reserved.