Diagnosis of mitochondrial disease: assessment of mitochondrial DNA heteroplasmy in blood.
Diagnosis of mitochondrial disease: assessment of mitochondrial DNA heteroplasmy in blood.
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DOI：
10.1006/bbrc.1998.9553
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发表时间：
1998-10
影响因子：
3.1
通讯作者：
R. Taylor;G. Taylor;C. Morris;J. Edwardson;D. Turnbull
中科院分区：
文献类型：
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作者：
R. Taylor;G. Taylor;C. Morris;J. Edwardson;D. Turnbull
线粒体DNA （mtDNA）突变是神经系统疾病的重要原因。鉴定致病mtDNA突变在血液中可能特别麻烦，因为通常存在低水平的突变mtDNA。最近的一项研究表明，细胞色素c氧化酶基因的异质性mtDNA突变被错误地认为与阿尔茨海默病有关。我们希望探讨血液mtDNA的分析是否会影响mtDNA疾病的诊断，这种分析是由许多DNA提取程序制备的。采用不同方法提取4例mtDNA异质突变患者的DNA，并采用放射性PCR-RFLP分析分析异质水平。虽然mtDNA异质性水平没有持续下降，但我们观察到，使用筛选tRNALeu(UUR) A3243G突变的引物，通过简单的“煮沸”程序提取的DNA样本中，出现了一种新的mtDNA假基因的共扩增。这个假基因很容易从rho度（无mtdna）细胞中提取的DNA中扩增出来，证实了它的核位置。因此，我们认为mtDNA假基因可能在准确鉴定血液中致病性异质mtDNA突变方面存在重大困难。
Mitochondrial DNA (mtDNA) mutations are an important cause of neurological disease. The identification of causative mtDNA mutations may be particularly troublesome in blood where there are often low levels of mutant mtDNA. This is evident from a recent study in which heteroplasmic mtDNA mutations in cytochrome c oxidase genes were incorrectly thought to be linked to Alzheimer's disease. We wished to explore whether analysis of blood mtDNA, prepared by a number of DNA extraction procedures, influenced the diagnosis of mtDNA disease. DNA was extracted by different procedures from 4 patients with heteroplasmic mtDNA mutations, and the level of heteroplasmy investigated by radioactive PCR-RFLP analysis. Whilst there was no consistent decrease in the level of mtDNA heteroplasmy, we observed the coamplification of a novel mtDNA pseudogene from DNA samples extracted by a simple 'boiling' procedure using primers designed to screen for the tRNALeu(UUR) A3243G mutation. This pseudogene was readily amplified from DNA extracted from rho degrees (mtDNA-less) cells, confirming its nuclear location. We believe that mtDNA pseudogenes may therefore present significant difficulties in the accurate identification of pathogenic heteroplasmic mtDNA mutations in blood.