MECHANISM OF DNA REPLICATION IN EUCARYOTES--II SV40 AS A MODEL SYSTEM

真核生物DNA复制机制--II SV40作为模型系统

• 批准号: 4693267
• 负责人: A SUGINO
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF ENVIRONMENTAL HEALTH SCIENCES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/4693267
• 关键词: DNA topoisomerases; Escherichia coli; antibody; bacteriophage M13; deoxyribonuclease I; enzyme structure; eukaryote; genetic models; human tissue; model design /development; monoclonal antibody; nucleic acid inhibitor; simian virus 40; virus DNA; virus genetics

DNA topoisomerases I and II, DNA primase, DNA-dependent ATPase and RNase H activities, all of which are expected to participate in DNA replication, have been purified to near homogeneity from human cell lines and calf thymus and monoclonal and polyclonal antibodies have been raised against each proteins. To determine whether these proteins are required for eucaryotic DNA replication, the soluble in vitro replication system of SV40 DNA (which mimics the in vivo system) has been established and used to test whether the antibodies inhibit in vitro SV40 DNA replication. This system makes possible not only the identification and purification of various DNA replication proteins but also the identification of other factors which maintain high fidelity of DNA synthesis. For measuring the fidelity of the in vitro SV40 replication system, chimeric M13mp2 phages have been constructed which consist of an SV40 DNA replication origin and the E. coli lacZ gene.

DNA拓扑异构酶I和II、DNA引发酶、DNA依赖性ATP酶和RNase H 活动，所有这些都有望参与DNA复制， 已从人细胞系和小牛中纯化至接近同质 胸腺和单克隆和多克隆抗体已经提出， 每个蛋白质。 为了确定这些蛋白质是否是 真核DNA复制，SV 40的可溶性体外复制系统 DNA（模拟体内系统）已经建立并用于测试 抗体是否抑制体外SV40 DNA复制。 该系统 不仅可以鉴定和纯化各种DNA， 复制蛋白质，但也确定其他因素， 保持DNA合成的高保真度。 用于测量 在体外SV40复制系统中，嵌合M13mp2质粒已被 构建了由SV40 DNA复制起点和E.杆菌 lacZ基因。