GENOMIC RESPONSES TO HORMONE SIGNALING

在任何给定的细胞中，只有所有编码基因的一个子集表达， 表达的水平和时间控制细胞发育， 分化、功能和生理学。在这里，我们描述了一种方法， 同时监测许多基因的表达水平， 并联该方法涉及测定相对 基于整个mRNA杂交的mRNA浓度 高密度合成寡核苷酸阵列 （基因芯片TM）。用于这里提出的实验的阵列 含有超过65，000种不同的25聚体寡核苷酸， 序列的面积为1.6平方厘米。在第一阶段，我们将：1）设计一套 的阵列（基因芯片TM），用于同时定量超过6500 人类基因（Hum 6000），来自GenBank和公共EST数据库（dbEST）。 2)证明该方法具有灵敏度、专属性和定量性。 3)使用Hum 6000芯片来确定由 三种激素受体（β-2肾上腺素能，CCR 5，催产素）代表 三种不同的信号通路在第二阶段，我们将使用悍马6000 芯片来分析由一个大的 一组受体和药理学探针。我们还将确认 通过使用北方分析， 蛋白质印迹和基因表达标记。 拟定商业应用： 定量监测大量的表达水平， 基因应该证明在阐明基因功能，探索 疾病的原因和机制，以及发现潜在的 治疗和诊断目标。作为基因组信息的主体 增长，这里描述的高度并行的方法提供了一个 利用序列信息来阐明 重要激素、药物和代谢途径。发展 基因芯片技术作为监测全基因组表达谱的工具 具有多种生物医学、制药和农业应用。 收入将来自基因芯片系统（射流）的销售 站点和扫描仪）和定制基因芯片研究与激素受体 将确定基因，这些基因将成为药物治疗的潜在新靶点， 发展脑内激活基因的调节区域可以 也可用于开发快速、非放射性的报告基因测定， 通过筛选激活激素受体的药物。

Only a subset of all encoded genes is expressed in any given cell, and the levels and timing of expression govern cellular development, differentiation, function and physiology. Here we describe a method for the simultaneous monitoring of the expression levels of many genes in parallel. The method involves the determination of the relative concentrations of mRNAs based on hybridization of entire mRNA populations to high-density arrays of synthetic oligonucleotides (GeneChips TM). The arrays used for the experiments proposed here contain more than 65,000 different 25-mer oligonucleotides of defined sequence in an area of 1.6 cm 2. In Phase I, we will: 1) Design a set of arrays (GeneChip TM) for the simultaneous quantitation of over 6500 human genes (Hum6000) from GenBank and the public EST database (dbEST). 2) Demonstrate that the method is sensitive, specific and quantitative. 3) Use the Hum6000 chip to determine the mRNAs dynamically regulated by three hormone receptors (beta-2 Adrenergic, CCR5, Oxytocin) representing three different signaling pathways. In Phase II we will use the Hum6000 chip to analyze the mRNAs that are dynamically regulated by a large panel of receptors, and pharmacologic probes. We will also confirm a subset of the findings from the Hum6000 chip by using Northern analysis, Western blots, and gene expression markers. PROPOSED COMMERCIAL APPLICATION: The quantitative monitoring of expression levels for large numbers of genes should prove valuable in elucidating gene function, exploring the causes and mechanisms of disease, and for the discovery of potential therapeutic and diagnostic targets. As the body of genomic information grows, highly parallel methods of the type described here provide an efficient and direct way to use sequence information to elucidate important hormone, drug and metabolic pathways. Development of the GeneChip technology as a tool to monitor genome-wide expression profiles has multifold biomedical, pharmaceutical and agricultural applications. Revenue will be derived from the sale of the GeneChip system (fluidics station and scanner) and custom GeneChips Studies with hormone receptors will identify genes that will be potential new targets for drug development. The regulatory regions of hormonally activated genes could also be used to develop rapid, non-radioactive reporter assays for high- throughput screening of drugs that activate hormone receptors.