GENETIC DISSECTION OF REPLICATION INITIATION IN YEAST

线粒体DNA复制启动机制的研究进展 人们对酿酒酵母知之甚少，部分原因是 高重组率使普通物理设备难以使用 接近了。然而，积累的证据表明，有两种机制 可能存在复制启动的。其一，类似于 哺乳动物线粒体DNA，可能利用启动子和转录裂解来产生 入门读物。另一种在不存在的情况下运行的机制 转录，可通过末端/内部启动复制分叉 重组事件。我们将使用一个组合来检查这些机制 结构和遗传方法。一种新的双向凝胶电泳法 技术将被用来系统地搜索线粒体基因组 用于各种可能的起爆结构。有温度的突变体 线粒体DNA复制启动中的敏感缺陷将被隔离 并以此为特征。转录和转录缺陷的双重突变体 将检查可能的重组途径是否存在维护故障 线粒体DNA。一种突变体，它废除了一类有偏见的遗传 线粒体缺失基因组，被认为具有复制优势 由于复制起始点高度集中，将被研究。

The mechanism of replication initiation of the mitochondrial DNA (mtDNA) in the yeast Saccharomyces cerevisiae is poorly understood, in part because a high rate of recombination has made it difficult to use common physical approaches. However, the accumulated evidence suggests that two mechanisms of replication initiation may exist. One, analogous to initiation in mammalian mtDNA, may employ a promoter and transcript cleavage to generate a primer. The other mechanism that operates in the absence of transcription, may initiate replication forks through end/internal recombination events. We will examine these mechanisms using a combination of structural and genetic approaches. A new 2-D gel electrophoresis technique will be used to systematically search the mitochondrial genome for various possible initiation structures. Mutants with a temperature sensitive defect in the initiation of mtDNA replication will be isolated and characterized. Double mutants defective in transcription and in possible recombination pathways will be examined for failure to maintain mtDNA. A mutant that abolishes the biased inheritance of a class of mitochondrial deletion genomes, thought to have a replication advantage because of a high concentration of replication origins, will be studied.