基于微阵列的嗅黏膜中高表达基因的发现[毕业论文].ppt

* * Microarray-based discovery of highly expressed olfactory mucosal genes: potential roles in the various functions of the olfactory system 基于微阵列的嗅黏膜中高表达基因的发现： 在嗅觉系统多功能作用中扮演的潜在角色 实验目的：获得在嗅黏膜中组织特异性基因表达的全局观。 OM呼吸道与神经系统的交界处，执行使两个器官系统连为一体的功能，是脊椎动物成体中神经发生和神经再生的主要位点。 嗅觉器官的感觉神经元负责气味转换，呼入的空气中有毒媒介能够被鼻呼吸道和嗅觉组织中的黏膜纤毛结构和代谢酶清除。OM相对于其它神经组织的独特之处在于：OM有一个基底细胞群，能够不断地提供新的神经元，使因受伤或毒气损伤而丢失的神经元得以替换。 MATERIALS AND METHODS SECTION 1 MATERIALS AND METHODS： 1、Animals, tissue samples, and RNA preparation 2、Fluorescent probe preparation and microarray hybridization 3、Date analyses 4、confirmatory test 5、Identi?cation of cis-elements in coordinately expressed genes 组织来源： 成鼠的嗅黏膜（OM）、嗅球（olfactory bulb）、鼻呼吸道黏膜（nasal respiratory mucosa）组织取自6-8周龄的C57BL/6雌性小鼠。 The ethmoid turbinates and caudal one-third of the nasal septum served as the source of OM RNA, and the nasoturbinates, maxillo-turbinates, and anterior two-thirds of the nasal septum were the source of respiratory mucosal RNA . Other tissues including multi-ple brain subregions, dorsal root ganglion, heart, lung, kidney, liver,reproductive and endocrine organs, immune organs, muscle, and skin are from a generalized mouse gene expression database. RNA提取： 用CO2吸入法处死小鼠 解剖小鼠，迅速取出组织，液氮速冻 将取自八只小鼠的组织在冰上匀浆 TRI Reagent提取总RNA 总RNA用酒精、醋酸钠沉淀 总RNA在DEPC水中溶解 Probe preparation and microarray hybridization were performed by Incyte Genomics (Palo Alto, CA) using the GEMBright random primer reverse transcription labeling kit (Incyte Genomics). Labeled cDNA was prepared from each poly(A)+ RNA sampleusing nucle- otides labeled with the ?uorescent dye Cy5. Labeled cDNAs were then subjected to competitive hybridization to mouse GEM1 microarrays, composed of spotted cDNAs corresponding to 8,638 sequence-veri?ed IMAGE expressed sequence tag (EST) cl