Liu V W, Zhang C, Nagley P
Department of Biochemistry and Molecular Biology, Monash University, Clayton, Victoria 3168, Australia.
Nucleic Acids Res. 1998 Mar 1;26(5):1268-75. doi: 10.1093/nar/26.5.1268.
In 60 human tissue samples (encompassing skeletal muscle, heart and kidney) obtained from subjects aged from under 1 to 90 years, we used quantitative PCR procedures to quantify mitochondrial DNA (mtDNA) molecules carrying the 4977 bp deletion (mtDNA4977) and 3243 A-->G base substitution. In addition, the prevalence of multiple mtDNA deletions was assessed in a semi-quantitative manner. For all three tissues, the correlations between the accumulation of the particular mtDNA mutations and age of the subject are highly significant. However, differential extents of accumulation of the two specific mutations in the various tissues were observed. Thus, the mean abundance (percentage of mutant mtDNA out of total mtDNA) of mtDNA4977in a subset of age-matched adults is substantially higher in skeletal muscle than in heart and kidney. However, the mean abundance of the 3243 A-->G mutation in skeletal muscle was found to be lower than that in heart and kidney. Visualisation of arrays of PCR products arising from multiple mtDNA deletions in DNA extracted from adult skeletal muscle, was readily made after 30 cycles of PCR. By contrast, in DNA extracted from adult heart or kidney, amplification for 35 cycles of PCR was required to detect multiple mtDNA deletions. Although such multiple deletions are less abundant in heart and kidney than in skeletal muscle, in all tissue extracts there are unique patterns of bands, even from different tissues of the same subject. The differential accumulation of mtDNA4977, other mtDNA deletions and the 3243 A-->G mutation in the three tissues analysed presumably reflects different metabolic and senescence characteristics of these various tissues.
在从1岁以下至90岁的受试者获取的60份人体组织样本（包括骨骼肌、心脏和肾脏）中，我们使用定量PCR程序对携带4977 bp缺失（mtDNA4977）和3243 A→G碱基替换的线粒体DNA（mtDNA）分子进行定量。此外，以半定量方式评估了多个mtDNA缺失的发生率。对于所有这三种组织，特定mtDNA突变的积累与受试者年龄之间的相关性非常显著。然而，观察到这两种特定突变在不同组织中的积累程度存在差异。因此，在一组年龄匹配的成年人中，mtDNA4977的平均丰度（突变型mtDNA占总mtDNA的百分比）在骨骼肌中显著高于心脏和肾脏。然而，发现骨骼肌中3243 A→G突变的平均丰度低于心脏和肾脏。在对从成年骨骼肌提取的DNA中多个mtDNA缺失产生的PCR产物阵列进行可视化时，在30个PCR循环后很容易实现。相比之下，在从成年心脏或肾脏提取的DNA中，需要进行35个PCR循环的扩增才能检测到多个mtDNA缺失。尽管这种多个缺失在心脏和肾脏中比在骨骼肌中少，但在所有组织提取物中都有独特的条带模式，即使是来自同一受试者的不同组织。在所分析的三种组织中，mtDNA4977、其他mtDNA缺失和3243 A→G突变的差异积累可能反映了这些不同组织的不同代谢和衰老特征。
