TESTS OF THE UNINEME MODEL FOR MAMMALIAN CHROMOSOME

哺乳动物染色体 UNINEME 模型的测试

• 批准号: 3302449
• 负责人: CHRISTOPHER S. LANGE
• 金额: $ 17.64万
• 依托单位: SUNY DOWNSTATE MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-07-01 至 1995-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3302449
• 关键词: DNA; blood viscosity; chemical structure function; chromosomes; flow cytometry; genetic models; human tissue; molecular genetics; nucleic acid structure; tissue /cell culture

This project will test the unineme hypothesis of chromosome structure in mammalian cells by a direct measurement of the molecular weight (Mr) of chromosomal DNA molecules in several mammalian species. Preliminary data, obtained by viscoelastometry (VE), for several species of rodent cells (V79 Chinese hamster lung, L5178Y mouse lymphoblastic leukemia and 9L rat brain tumor) show that mammalian (or at least rodent) DNA molecules have an Mr equal to only 1/4 to 1/8 the average chromatid DNA content. This implies a multimolecular DNA chromosomal architecture. A second preliminary observation in disagreement with the unineme hypothesis is that rodent DNA molecules increase their hydrodynamic (excluded) volume when irradiated with ionizing radiation doses which would be expected to create an average of one double-strand break (DSB) per molecule. Such behavior is expected for circular molecules. No data exist for human cells to determine if they are the sam or different in this regard. However, the different dose dependence for the creation of prematurely condensed chromosome (PCC) fragments suggests that human cells may have linear DNAs. Therefore, data will be obtained to determine if rodent and human chromosomal DNA molecules are linear or circular and their size(s). Two approaches will be used. First, DNA size and shape will be determined on cell lysates; this will yield the size and shape of the largest DNA molecules. Studies will be performed to determine the sensitivity of VE to detect size heterogeneity so that we can determine if all DNA molecules in a nucleus are the same size or different. The second approach will be to sort specific large and small chromosomes by flow cytometry and measure the size (and shape) of their DNAs by VE. Subsidiary studies will be performed to increase the accuracy of DNA size determination in the high ionic strength solutions used to deproteinize the DNA, and to control for any effects of the dyes used for flow sorting of the chromosomes. Thus, these studies will: 1) determine the validity of the unineme model for rodent and human cells, 2) determine the size and shape of the DNA molecules of rodent and human chromosomes, and 3) determine the size of the DNA molecules of specific human and rodent chromosomes.

本计画将检验染色体结构之单丝体假说， 哺乳动物细胞的分子量（Mr）的直接测量 几种哺乳动物的染色体DNA分子。 初步数据， 通过粘弹性测定法（VE）获得，用于几种啮齿动物细胞（V79 中国仓鼠肺、L5178 Y小鼠淋巴细胞白血病和9 L大鼠脑 肿瘤）表明哺乳动物（或至少啮齿动物）DNA分子具有MR 仅相当于平均染色单体DNA含量的1/4至1/8。 这意味着 多分子DNA染色体结构。 第二初步 与单丝体假说不一致观察结果是， 分子增加其流体动力学（排除）体积时，照射 电离辐射剂量，预计将产生平均 一个双链断裂（DSB）。 这样的行为是意料之中的 for circular圆形molecules分子. 目前还没有关于人类细胞的数据来确定它们是否 在这方面是相同的还是不同的。 然而，不同的剂量 产生过早凝聚染色体的依赖性 片段表明人类细胞可能具有线性DNA。 因此数据 以确定啮齿动物和人类的染色体DNA分子 是线性的或圆形的，以及它们的大小。 将采用两种方法。 首先，将在细胞裂解物上确定DNA大小和形状;这将 产生最大DNA分子的大小和形状。 研究将 测定VE检测大小异质性的灵敏度 这样我们就可以确定细胞核中的DNA分子是否都是一样的 大小或不同。 第二种方法是对特定的大型和 通过流式细胞术测量小染色体的大小（和形状）， 他们的DNA 将进行辅助研究，以增加 高离子强度溶液中DNA大小测定的准确性 用来去除DNA的蛋白质，并控制染料的任何影响， 用于染色体的流式分选。 因此，这些研究将：1） 确定啮齿动物和人类细胞的unineme模型的有效性，2） 确定啮齿动物和人类DNA分子的大小和形状 染色体，和3）确定特定的DNA分子的大小 人类和啮齿动物的染色体。