Development of a Genome-wide Drosophia RNAi Resource

描述（由申请人提供）：遗传学为解剖生物功能提供了最具洞察力和最有力的方法之一，果蝇已成为此类分析的主要模式生物，其中许多已被探索与人类健康相关的过程。现在，RNAi的基因失活正在彻底改变遗传方法。通过使极快的全基因组筛选成为可能，全基因组果蝇RNAi （dmRNAi）资源将极大地加快果蝇的遗传研究。RNAi方法将立即建立功能和分子数据之间的联系。此外，通过为这些分析建立一个双链rna的参考集合，在许多研究不同问题的实验室收集的数据可以组装成一个共同的功能数据库，这将鼓励发现新的连接。在成功构建果蝇大多数保守基因双链rna不完整文库的基础上，我们将首先建立一个完整的文库。然后，我们将开发降低成本、促进分销、鼓励使用和增加方法灵活性的方法。我们将探索以可接受的成本结构在全基因组范围内制备RNA的方法。最后，将探索一种标记策略，作为一种方法，通过允许DNA模板池来压缩复杂的文库，从而可以从5个试管中存储和检索14,000个构建的文库。文库本身和提出的技术进步将通过提供覆盖整个果蝇基因组的完整集合，促进数据积累到公共数据库，降低这些研究的成本，并最大限度地减少启动全基因组RNAi筛选的技术障碍，从而使RNAi基因失活的能力广泛可用。三种产品将在这笔资金下商业化：(1)代表所有果蝇基因的DNA构建物用于产生RNA，(2)来自RNAi“就绪”的DNA构建物的RNA，以及(3)在基因组标记策略下可用的一系列池，用于筛选整个基因组。

DESCRIPTION (provided by applicant): Genetics has provided one of the most discerning and powerful approaches to the dissection of biological function and Drosophila has served as a major model organism for such analyses, many of which have been probed processes relevant to human health. Now gene inactivation by RNAi is revolutionizing genetic approaches. By making possible extremely rapid whole genome screens, a Genome-wide Drosophila RNAi (dmRNAi) resource will tremendously accelerate genetic studies in Drosophila. The RNAi approach will immediately establish a link between function and molecular data. Furthermore, by establishing a reference collection of double stranded RNAs for these analyses, data gathered in numerous laboratories studying diverse problems can be assembled into a common functional data base that will encourage discovery of novel connections. Building on successful construction of an incomplete library of double stranded RNAs representing most conserved genes of Drosophila, we will first establish a complete library. We will then develop approaches to reduce cost, facilitate distribution, encourage use and increase the flexibility of the methods. We will explore methods for the preparation of RNA on a genome-wide scale at an accessible cost structure. Finally, a tagging strategy will be explored as an approach that should allow compression of complex libraries by allowing pooling of DNA templates such that a library of 14,000 constructs can be stored and retrieved from 5 tubes. The library itself and the proposed technical advances will lead the way in making the power of gene inactivation by RNAi available widely by providing a complete collection covering the entire Drosophila genome, facilitating the accrual of data into a common data base, reducing the cost of these studies, and minimizing the technical impediments to initiating genome-wide RNAi screens. Three products will be commercialized under this funding: (1) DNA constructs representing all of the Drosophila genes for use in generating RNA, (2) RNA from the DNA constructs that are RNAi 'ready', and (3) a series of pools available under the genome tagging strategy useful in screening the entire genome.