NUCLEAR AND MITOCHONDRIAL DNA FROM WILD-TYPE AND PETITE YEAST - CIRCULARITY, LENGTH, AND BUOYANT DENSITY
NUCLEAR AND MITOCHONDRIAL DNA FROM WILD-TYPE AND PETITE YEAST - CIRCULARITY, LENGTH, AND BUOYANT DENSITY
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DOI：
10.1073/pnas.64.2.739
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发表时间：
1969-01-01
影响因子：
11.1
通讯作者：
AVERS, CJ
中科院分区：
文献类型：
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作者：
BILLHEIMER, FE;AVERS, CJ
纯化的线粒体和核DNA从二倍体同基因野生型和植物娇小的面包酵母进行了分析，通过电子显微镜和CsCl梯度的分析ultracentrugation。核DNA在氯化铯中的浮力密度在两个菌株中相同（ρ = 1.700），但线粒体DNA在野生型（ρ = 1.684）和娇小型（ρ = 1.680）之间存在差异。电子显微镜下发现两个菌株的细胞核和线粒体DNA的圆形和线性丝。核DNA分子包括6.5%的环状细丝，主要测量轮廓长度为2 μ或更小，和线性细丝，显示单峰，分散的长度分布集中在约2至3 μ，对于两个菌株。野生型的线粒体DNA因提取和纯化分子的方法而异; CsCl亚分级样品中只有7.5%的环状分子，而氯仿提取的DNA中没有经过CsCl处理的环状分子为15%，而在早期的研究中，线粒体裂解的环状分子高达50%。圆的模态长度出现在约2、5和10 μ处。在使用三种制备方法的线性分子的长度分布模式的比较中，增加的剪切降解也是明显的。小线粒体DNA含有36- 38%的环状分子，其测量值为0.3-5.3 μ，但主要在0.3 - 2.0 μ的范围内，无论是来自氯仿提取的群体还是来自CsCl亚分级的群体。先前对线粒体裂解的研究显示最多有8%的圆圈，我们现在将其归因于当时未能检测到小于1 μ的圆圈，这是本研究中纯化DNA样品中遇到的重要成分。线性细丝在每种情况下呈现单峰长度distribution.尽管来自三种不同的制备方法的分子群体的变化，有一致的差异线粒体DNA从野生型和小酵母的频率和大小的环状分子，以及长度分布模式。
Purified mitochondrial and nuclear DNA from diploid isogenic wild-type and vegetative-petite baker's yeast were analyzed by electron microscopy and by analytical ultracentrifugation in CsCl gradients. The buoyant densities in CsCl of nuclear DNA were identical for the two strains (ρ = 1.700), but there was a difference between mitochondrial DNA from the wild type (ρ = 1.684) and the petite (ρ = 1.680). Electron microscopy revealed both circular and linear filaments for nuclear and for mitochondrial DNA of both strains. Nuclear DNA molecules included 6.5 per cent cyclic filaments principally measuring 2 μ or less in contour length, and linear filaments showing a unimodal, disperse length-distribution centered at about 2 to 3 μ, for both strains. Mitochondrial DNA for wild type varied depending upon the method used to extract and purify the molecules; showing only 7.5 per cent circular molecules from CsCl-subfractionated samples, as compared with 15 per cent circles from chloroform-extracted DNA not subjected to CsCl and up to 50 per cent circles from osmotically-lysed mitochondira, as reported in an earlier study. Modal lengths of circles occurred at about 2, 5, and 10 μ Increasing shear degradation also was evident in comparisons of the length-distribution patterns of linear molecules using the three preparative methods. Petite mitochondrial DNA contained 36-38 per cent circular molecules which measured 0.3-5.3 μ, but principally in the range of 0.3 to 2.0 μ whether from chloroform-extracted populations or from ones subfractionated in CsCl. A previous study of osmotically lysed mitochondria had shown a maximum of 8 per cent circles, which we now attribute to a failure, at that time, to detect circles measuring less than 1 μ, a substantial component encountered in the purified DNA samples in the present study. Linear filaments presented a unimodal length distribution in every case.Despite the variation in molecule populations derived from the three different preparative methods, there were consistent differences between mitochondrial DNA from wild-type and petite yeast in frequencies and size of circular molecules, as well as in length distribution patterns.