Molecular Typing of Taste Cells Using Microarrays

描述(申请人提供)：长期目标是了解 不同类型的细胞形态和形态的分子机制 生理异质味觉细胞在接受、加工和 传送味觉信号。利用一组已知的基因，我们已经成功地 应用单细胞RT-PCR扩增和筛选杂交方法 确定一些味觉细胞的有限基因表达模式，并识别 几种味觉细胞类型的选择性信号元件被认为起着关键的作用 在苦涩的感觉中扮演的角色。在这个应用中，我们将使用单细胞RT-PCR 探测DNA阵列以建立全球基因表达谱的产品 许多味觉细胞，以确定和预测味觉细胞类型/亚型基于 它们的表达特征，以识别额外的口味类型或亚型- 决定这一类型或亚型的选择性基因或基因簇 味觉的生理功能。这一行动的具体目标 应用：1.从味蕾、丝状乳头中产生DNA探针 和100个单独的味觉细胞同时使用Arna扩增方法和 改良的单细胞RT-PCR程序；2.筛选全基因组DNA阵列 味蕾和丝状探针，以识别和重新排列 在丝状乳头上富含味蕾，以制备味觉DNA阵列(味觉 薯片)；3.用味蕾探头成对筛选味觉芯片 与要产生的100个单一味觉细胞探针之一配对，并 通过以下方式建立个体味觉细胞的基因表达谱 在每个单细胞探针中识别味觉芯片上表达的基因，以 使用非监督工具确定和发现味觉细胞类型或亚型 分区聚类法，并用来识别其他基因分类器 邻域分析方法。这些研究的结果将有助于 后基因组生物医学新技术的发展 研究，为味觉细胞分类和产量提供遗传基础 对味觉潜在分子机制的重大新见解 转导。从这项应用中获得的知识应该会进一步推动我们的 理解味觉敏感度的遗传差异，以及味觉和 代谢性疾病，如厌食症、厌食症和恶病质。

DESCRIPTION (provided by applicant): The long-term objective is to understand the molecular mechanisms that different types of morphologically and physiologically heterogeneous taste cells utilize in receiving, processing and transmitting gustatory signals. Using a set of known genes we have successfully applied single cell RT-PCR amplification and filter hybridization methods to determine limited gene expression patterns for some taste cells, and to identify several taste cell type selective signaling elements thought to play a critical role in bitter sensation. In this application, we will use single cell RT-PCR products to probe DNA arrays to establish global gene expression profiles for many taste cells, to determine and predict taste cell types/subtypes based on their expression profiles, to identify additional taste type or subtype- selective genes or clusters of genes that define this type or subtype's physiological functions in taste perception. The specific goals of this application are: 1. To generate DNA probes from taste buds, filiform papillae and 100 individual taste cells using both an aRNA amplification method and a modified single cell RT-PCR procedure; 2. To screen genome-wide DNA arrays with both taste bud and filiform probes, to identify and re-array genes that are enriched in taste buds over filiform papillae to prepare taste DNA arrays (taste chips); 3. To screen the taste chips in pair wise fashion with a taste bud probe paired with one of the 100 single taste cell probes to be generated, and to establish the gene expression profiles of the individual taste cells by identifying the expressed genes on the taste chips in each single cell probe, to determine and discover taste cell types or subtypes using unsupervised partitional clustering method, and to identify additional gene classifiers with the neighborhood analysis method. The results of these studies will contribute to the development of new technologies in pursuing post-genomic biomedical research, provide genetic bases for taste cell classification and yield significant novel insights into the molecular mechanisms underlying taste transduction. The knowledge gained from this application should further our understanding of genetic differences in taste sensitivity, and the gustatory and metabolic disorders such as malgeusia, dysgeusia and cachexia.