STUDIES OF BACTERIOPHAGE MU DNA TRANSPOSITION & EXCISION

我们建议研究DNA转座的某些特定方面，并 噬菌体Mu中的切除。我们想要跟进我们的惊喜 观察到Mu的移位可以通过两条途径发生，并且 影响路径选择的因素之一可能是 所需的两种蛋白质A和B的比例 换位。我们将利用这一观察结果来具体研究一个 或者另一条路。除了参与DNA转座外，A 蛋白质还导致Mu DNA序列从它们的 综合站点。我们已经在强LacUV5下克隆了A基因 推动者。在过量生产A的情况下，我们可以在体内检测到线性切除 MU DNA。我们建议利用这一观察结果来测试切除的Mu DNA 是换位过程中的中间体。我们还计划复制 在体外发生切除反应，从而有一种纯化A蛋白的方法。 最终我们计划在足够的条件下获得纯转座蛋白 研究蛋白质和蛋白质之间的物理化学相互作用 它们的底物。

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.