GENETIC ANALYSIS OF MAMMALIAN MITOCHONDRIAL INHERITANCE

哺乳动物线粒体DNA转移机制的细节 (MtDNA)从母亲传给后代以及这 线粒体遗传多样性起源的机制 目前还不得而知。线粒体DNA序列的频繁出现 物种内的多态表明，一个物种的简单转移 雌性的线粒体对其后代的补充没有发生，并且 遵循一条更复杂的路径，该路径仅传输一小部分 从母体线粒体转移到新胚胎。通过这条路 瓶颈将允许新线粒体的快速出现 人群中的基因分型。 为了测试这个模型，异质小鼠(它包含两个不同的 MtDNA基因组)是通过在单细胞上进行胚胎融合而构建的 水平。对异质雌性小鼠后代的分析表明，两者都 线粒体DNA物种被遗传给后代；他们似乎是 在不同的后代中以不同的水平传播。 描述了将确定分离率是否为 在核基因背景不同的两个线粒体DNA物种中 完全相同，以及总体线粒体DNA异质性和 异质性水平可能存在组织差异。在……里面 此外，两个不同线粒体DNA物种的分离速度 从这些动物衍生的细胞系中将被确定。最后，一个 描述了一种检测父亲线粒体DNA的敏感试验。这些 实验将有助于确定线粒体DNA变异的种类和数量 这可能发生在哺乳动物身上。 这项提案的第二个主要目标是开发检测线粒体DNA的方法。 转型。来自不同来源的线粒体将被转移 从不同类型的体细胞转化为胚胎。一旦这是 完成后，将尝试将mtDNA导入细胞器 使用化学处理、脂质体融合、电穿孔或使用 粒子加速器用于转化细胞系或分离的 线粒体上有外源mtDNA。这些实验将允许 体外携带突变线粒体DNA的动物的构建，并允许 线粒体DNA功能在肌肉和神经中作用的研究 肌病。这些动物和让它们的 构筑也应该有助于理解细胞质运动 在早期胚胎发育期间，细胞谱系的移动，以及 线粒体在早期发育过程中发生的变化。

The details of the mechanism which transfers mammalian mitochondrial DNA (mtDNA) from mother to offspring and the constraints which this mechanism would place on the origin of mitochondrial genetic diversity are currently unknown. The frequent occurrence of mtDNA sequence polymorphisms within a species suggests that simple transfer of a female's mitochondrial complement to her offspring does not occur and that a more complex path is followed which transfers only a small subset of the maternal mitochondria to the new embryo. Passage through this bottleneck would allow for the rapid appearance of new mitochondrial genotypes in the population. To test this model, heteroplasmic mice (which contain two different mtDNA genomes) were constructed by embryo fusion at the single cell level. Analysis of offspring of heteroplasmic female mice indicate both mtDNA species are transmitted to offspring; they appear to be transmitted at different levels in different progeny. Experiments are described which will determine if the segregation rates of the two mtDNA species in different nuclear gene backgrounds are identical, and whether both the level of overall mtDNA heteroplasmy and possible tissue differences in the levels of heteroplasmy occur. In addition, the rates at which the two different mtDNA species segregate in cell lines derived from these animals will be determined. Finally, a sensitive test for the presence of paternal mtDNA is described. These experiments will help define the kind and amounts of mtDNA variation that can occur in mammals. A second major goal of this proposal is to develop methods for mtDNA transformation. Mitochondria from various sources will be transferred into embryos, from somatic cells of various types. Once this is accomplished, mtDNA introduction into the organelle will be attempted using chemical treatment, liposome fusion, electroporation, or use of a particle accelerator to transform either cell lines or isolated mitochondria with exogenous mtDNA. These experiments will allow the construction of animals carrying in vitro mutated mtDNAs, and permit an examination of the role of mtDNA function in muscle and nerve myopathies. These animals and the techniques that allow their construction should also help in understanding cytoplasmic movement during early embryo development, movement of cell lineages, and the changes in mitochondria which occur during early development.