IDENTIFICATION OF GENES INDUCED OR REPRESSED BY CHRONIC ALCOHOL

该项目提出了一种新的方法来识别基因， 在调节小鼠酒精戒断的严重程度的重要性， 鉴定慢性乙醇暴露过程中表达和调节的mRNA。 潜在的假设是，基因表达的变化很可能 在大脑中发生慢性暴露于乙醇后， 其表达受到慢性乙醇的影响是一个很好的候选者， 在乙醇神经适应中具有重要的功能作用。 基因 受慢性乙醇暴露的调节可能会增加戒断风险，或 防止撤回的专家保护。 我们建议识别新的 候选基因的重要性，没有偏见的潜在作用，使用 差异显示技术。 将通过吸入使近交WSP-2小鼠产生身体依赖性，并且mRNA 将通过以下方法分析暴露小鼠和对照小鼠脑中的 差异显示 依赖性和对照RNA将与 修饰的oligo-dT引物并逆转录。 所述逆转录的 cDNA序列将用随机引物子集通过PCR扩增， 标记产物并在DNA测序凝胶上分离。 通过比较 不同群体的带型， 将忽略对照和处理样品中的强度， 留下许多差异展示的基因产物（即，要么 上调或下调），可能是乙醇对 基因表达。

The proposed project takes a novel approach to identifying the genes of importance in modulating the severity of alcohol withdrawal in mice by identifying mRNAs expressed and regulated during chronic ethanol exposure. The underlying hypothesis is that changes in gene expression are likely to occur in the brain after chronic exposure to ethanol, and that any gene whose expression is affected by chronic ethanol is a good candidate for having a functionally important role in ethanol neuroadaptation. Genes regulated by chronic ethanol exposure might promote withdrawal risk, or expert protection against withdrawal. We propose to identify novel candidate genes of importance, without bias about potential roles, using the technique of differential display. Inbred WSP-2 mice will be made physically dependent by inhalation, and mRNA from the brains of exposed and control mice will be analyzed by differential display. Dependent and control RNAs will be hybridized to modified oligo-dT primers and reverse transcribed. The reverse transcribed cDNA sequences will be amplified by PCR with subsets of random primers, and the products labeled and separated on a DNA sequencing gel. By comparing the banding patterns for the different groups, bands with similar intensities in both the control and treated samples will be ignored, leaving a number of gene products differentially displayed (i.e., either up- or down-regulated) presumably as a result of an effect of ethanol on gene expression.