Molecular Typing of Taste Cells Using Microarrays

描述（由申请人提供）：长期目标是了解 不同类型的形态和 生理上异质的味觉细胞用于接收、处理和 传递味觉信号利用一组已知的基因，我们成功地 应用单细胞RT-PCR扩增和滤膜杂交方法， 确定某些味觉细胞的有限基因表达模式，并鉴定 几种味觉细胞类型选择性信号元件被认为在 在苦味中的作用。在本申请中，我们将使用单细胞RT-PCR 产品用于探测DNA阵列以建立全球基因表达谱， 许多味觉细胞，以确定和预测味觉细胞类型/亚型，基于 他们的表达谱，以确定其他口味类型或亚型- 定义这种类型或亚型的选择性基因或基因簇 味觉的生理功能。具体目标是 应用是：1.为了从味蕾丝状乳头中提取DNA探针 和100个单独的味觉细胞，使用aRNA扩增方法和 改良的单细胞RT-PCR方法; 2.筛选全基因组DNA阵列， 味蕾和丝状探针，以确定和重新排列基因， 在丝状乳头上的味蕾中富集以制备味觉DNA阵列（味觉 芯片）; 3.用味蕾探针成对筛选味觉芯片 与待产生的100个单味细胞探针之一配对， 通过以下步骤建立单个味觉细胞的基因表达谱： 在每个单细胞探针中鉴定味觉芯片上表达的基因， 确定和发现味觉细胞类型或亚型， 分区聚类方法，并确定额外的基因分类器， 邻域分析法这些研究的结果将有助于 新技术的发展，追求后基因组生物医学 研究，为味细胞分类和产量提供遗传学基础 对味觉潜在分子机制的重要新见解 转导从这个应用程序中获得的知识应该进一步促进我们的 理解味觉敏感性的遗传差异，味觉和 代谢紊乱，如味觉缺失、味觉障碍和恶病质。

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.