DNA SEQUENCING OF UV-INDUCED CHROMOSOMAL MUTATIONS

紫外线诱导的染色体突变的 DNA 测序

• 批准号: 3282091
• 负责人: Christopher W Lawrence
• 金额: $ 13.74万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 1983
• 资助国家: 美国
• 起止时间: 1983-12-01 至 1991-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3282091
• 关键词: DNA repair; Escherichia coli; bacterial DNA; bacterial genetics; chemical addition; chromosome aberrations; gene deletion mutation; genetic manipulation; genetic mapping; genetic recombination; lac operon; nucleic acid sequence; plasmids; pyrimidines; radiation genetics; regulatory gene; synthetic nucleotide; ultraviolet radiation

Our objective is to understand the molecular mechanisms responsible for misrepair mutagenesis in Escherichia coli. These processes are elicited in response to DNA damage inflicted by a variety of mutagens, both physical and chemical, that occur in our environment naturally or are produced by human activity. These agents not only cause mutations, but, by altering certain regulatory genes, may also play a role in carcinogenesis. Such mutagens are a potential health hazard, therefore, though the extent to which this potential for harm is realized is not yet clear in many cases. Direct measurement of hazard at the low levels of damage usually encountered is rarely feasible, and extrapolation from experimental results with high doses of mutagen administered to non-human organisms is hampered by lack of information about the relevant mechanisms. Work in this project should help to provide such information. Our specific goals are to determine the types and frequencies of mutations induced by specific DNA lesions, namely, cyclobutane bipyrimidines and 6,4 Pyo adducts; to obtain evidence in vivo and in vitro for bypass replication, the process thought to cause mutations, and to see if the mechanisms for bypass replication differ when single- as opposed to double-stranded DNA is replicated; to investigate the properties of the umuDC gene, which is thought to play an important role in bypass replication; and to investigate the phenomenon of untargeted mutagenesis as a means of gaining insight into changes in the replication complex that result from UV irradiation. We will use a combination of genetical and molecular biological techniques to achieve these aims, including the sequencing of induced mutations, the construction of synthetic oligonucleotide hybrid phages containing a single defined lesion at a specific site, and the development of an in vitro system with which to study bypass replication.

我们的目标是了解 大肠杆菌中的错误修复诱变。 这些过程是在 对各种诱变剂造成的DNA损伤的反应，包括物理诱变剂和化学诱变剂。 和化学物质，它们在我们的环境中自然产生， 人类活动。 这些因子不仅会引起突变， 某些调控基因也可能在致癌作用中发挥作用。 等 诱变剂是一种潜在的健康危害，因此，虽然程度， 在许多情况下，这种潜在的伤害是如何实现的尚不清楚。 在低损害水平上直接测量危害通常 遇到的是很少可行的，从实验结果推断 对非人类生物体施用高剂量的诱变剂 缺乏有关机制的信息。 在这个项目中工作 应该有助于提供这种信息。 我们的具体目标是确定突变的类型和频率 诱导的特定DNA损伤，即环丁烷双嘧啶和6，4 Pyo加合物;在体内和体外获得旁路的证据 复制，这一过程被认为会导致突变，并观察 旁路复制的机制在单个复制时不同， 复制双链DNA;为了研究双链DNA的性质， umuDC基因，被认为在旁路中起重要作用， 复制;并研究非靶向诱变的现象， 一种深入了解复制复合体中的更改的方法， 紫外线照射的结果。 我们将结合遗传学和分子生物学技术 为了实现这些目标，包括诱导突变的测序， 含有单一寡核苷酸的合成寡核苷酸杂合引物的构建 特定部位的明确病变，以及体外 研究旁路复制的系统。