STUDIES OF BACTERIOPHAGE MU DNA TRANSPOSITION & EXCISION

噬菌体MU DNA转座的研究

• 批准号: 3282699
• 负责人: Rasika M Harshey
• 金额: $ 3.38万
• 依托单位: SCRIPPS CLINIC AND RESEARCH FOUNDATION
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-04-01 至 1988-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3282699
• 关键词: Escherichia coli; bacterial virus; chromosome deletion; chromosome translocation; genetic manipulation; genetic mapping; plasmids; virus DNA

We propose to investigate certain specific aspects of DNA transposition and excision in bacteriophage Mu. We would like to follow up our surprising observation that transposition of Mu can occur via two pathways and that one of the factors that influence the choice of the pathways could be the proportion of the two proteins A and B that are required for transposition. We will exploit this observation to study specifically one or the other pathway. Besides its involvement in DNA transposition, the A protein also brings about the excision of Mu DNA sequences from their integrated sites. We have cloned the A gene under the strong lacUV5 promoter. Upon overproduction of A, we can detect in vivo, linear excised Mu DNA. We propose to use this observation to test whether excised Mu DNA is an intermediate in transposition. We also plan to reproduce the excision reaction in vitro and thus have an assay to purify the A protein. Eventually we plan to obtain pure transposition proteins in sufficient amounts to study the physico-chemical interactions between the proteins and their substrates.

我们建议研究DNA转座的某些特定方面， 噬菌体Mu中的切除。 我们想继续我们令人惊讶的 观察到Mu的转位可以通过两种途径发生， 影响途径选择的因素之一可能是 两种蛋白质A和B的比例， 换位 我们将利用这一观察来具体研究一个 或者另一条路。 除了参与DNA转位，A 蛋白质也带来了Mu DNA序列的切除， 综合网站。 我们克隆了强lacUV5下的A基因， 启动子 当A过量产生时，我们可以在体内检测线性切除的 Mu DNA。 我们建议利用这一观察结果来测试是否切除的Mu DNA 是转座过程中的一个中间体。 我们还计划复制 在体外进行切除反应，从而具有纯化A蛋白的测定。 最终，我们计划在足够的条件下获得纯的转座蛋白。 相当于研究蛋白质之间的物理化学相互作用， 它们的基质。