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目的探讨EB病毒转化形成的永生细胞对线粒体DNA突变比例及拷贝数的影响。方法随机选择23位携带线粒体A3243G突变的患者,采集静脉血7ml,其中2ml提取总DNA,为对照组;5ml用EB病毒转化成为永生细胞,提取总DNA,为实验组。采用Taqman探针联合扩增受阻突变体系定量聚合酶链反应(ARMS-q PCR)方法检测线粒体A3243G突变比例和拷贝数。运用配对t检验进行统计分析。结果实验组的线粒体A3243G突变比例与对照组无明显差异;实验组的野生型线粒体DNA拷贝数,突变型线粒体DNA拷贝数和总线粒体DNA拷贝数与对照组比较,均无明显差异。结论 EB病毒转化对线粒体DNA数量上无明显影响,证实了通过EB病毒转化建立的永生细胞株可以长久保存线粒体DNA,为线粒体相关疾病的研究提供充足的实验材料。
Objective To investigate the effect of immortalized cells transformed by Epstein-Barr virus (EBV) on mitochondrial DNA mutation ratio and copy number. Methods Twenty-three patients with mitochondrial A3243G mutation were randomly selected, and 7ml venous blood was collected, of which 2ml total DNA was extracted and used as the control group. 5ml EB virus was transformed into immortalized cells and the total DNA was extracted for the experimental group. Taqman probe combined with amplification-blocked mutant was used to detect mitochondrial A3243G mutation ratio and copy number by quantitative polymerase chain reaction (ARMS-q PCR). Using paired t test for statistical analysis. Results The mitochondrial A3243G mutation rate in the experimental group was not significantly different from that in the control group. There was no significant difference in the copy number of the wild-type mitochondrial DNA, the copy number of the mutant mitochondrial DNA and the total copy number of the mitochondrial DNA in the experimental group compared with the control group. Conclusion EB virus transformation has no significant effect on the amount of mitochondrial DNA, confirming that the immortalized cell line established by EB virus transformation can preserve mitochondrial DNA for a long time and provide sufficient experimental materials for the study of mitochondria-related diseases.