EXPRESSION OF AN INTERRUPTED PROKARYOTIC GENE

这项研究的长期目标是利用TD基因的分裂， 噬菌体T4作为理解RNA和DNA的分子范例 自剪接的移动的内含子的交易。在第一个5年期间， 资助期间（由NSF资助DMB 8502961支持），我们确定， td间插序列是I组自剪接内含子。我们用这个 检测T4中的另外两个内含子的知识，并发现其中一个 这些和TD内含子是移动的，有效地转移到 它们各自基因的无内含子等位基因。至于它们的真核生物群 I对应物，内含子移动性依赖于编码的核酸内切酶。 由内含子本身决定。拟议的研究旨在进一步促进我们的 对基于RNA（剪接）和 基于DNA的（迁移）过程，使用td基因作为模型系统。对 我们将继续利用强大的积极和消极的 由该噬菌体遗传系统和TD基因提供的选择， 特别的。对于剪接研究，我们将继续使用分子标记。 遗传学来分析原核核酶的功能。我们将 进行平行研究，以调查大肠杆菌的可能作用 大肠杆菌作为自我剪接过程的附属物。相对于 DNA迁移率，我们将检查td核酸内切酶的功能，它 介导这一过程，以及它与其DNA靶标内含子的相互作用 归航地点我们还将问一些关于 内含子-噬菌体关系，并使用td基因作为探索 原核系统中内含子丢失的可能性。最后，合理化 内含子的存在，在流线型噬菌体基因组，我们希望解决 内含子通过增强选择性优势的假设 噬菌体重组性总之，拟议的实验代表了一个 继续进行正在进行的工作，利用简单的原核遗传 技术来阐明I组内含子的多方面反应， 它们是原核生物和真核生物共有的动态遗传元件， 王国。

The long-term objective of this research is to use the split td gene of phage T4 as the molecular paradigm for understanding the RNA and DNA transactions of a self-splicing, mobile intron. During the first 5-year funding period (supported by NSF grant DMB8502961) we established that the td intervening sequence is a group I self-splicing intron. We used this knowledge to detect two other introns in T4 and discovered that one of these as well as the td intron are mobile, transferring efficiently to intronless alleles of their respective genes. As for their eukaryotic group I counterparts, intron mobility is dependent upon an endonuclease encoded by the intron itself. The proposed research is intended to further our fundamental understanding of aspects of both the RNA-based (splicing) and DNA-based (mobility) processes, using the td gene as model system. To both ends we shall continue to exploit the powerful positive and negative selections provided by this phage genetic system and by the td gene in particular. For the splicing studies we shall continue to use molecular genetics to analyze the function of a prokaryotic ribozyme. We shall conduct parallel studies to investigate the possible role of Escherichia coli functions as accessories to the self-splicing process. With respect to DNA mobility, we shall examine the function of the td endonuclease, which mediates the process, and its interaction with its DNA target, the intron homing site. We shall also ask some more general questions about the intron-phage relationship and use the td gene as a model for exploring the possibility of intron loss in a prokaryotic system. Finally, to rationalize the existence of introns , in streamlined phage genomes, we wish to address the hypothesis that introns provide a selective advantage by enhancing phage recombinogenicity. Together, the proposed experiments represent a continuation of ongoing work that exploits facile prokaryotic genetic techniques to shed light on the multifaceted reactions of group I introns, which are dynamic genetic elements in common to the pro- and eukaryotic kingdoms.