Identification of Candidate Genes for Alcohol Preference by Expression Profiling of Congenic Strains. Rattus norvegicus

A highly significant quantitative trait locus (QTL) that influenced alcohol preference was identified in the iP/iNP rats on chromosome 4. Congenic strains in which the iP chromosome 4 QTL interval was transferred to the iNP (NP.P) exhibited the expected increase in alcohol consumption compared to the iNP background strain. This study was undertaken to identify genes in the chromosome 4 QTL interval that might contribute to the differences in alcohol consumption between the alcohol-naïve congenic and background strains. RNA from five brain regions from each of 6 NP.P and 6 iNP rats was labeled and analyzed separately on an Affymetrix Rat Genome 230 2.0 microarray. Expression levels were normalized using robust multi-chip average (RMA). Differential gene expression was validated using quantitative real-time PCR. An analysis combining five brain regions, including nucleus accumbens, frontal cortex, amygdala, hippocampus, and striatum, identified twenty three transcripts and nine ESTs that were differentially expressed between the NP.P and iNP. Of the twenty three observed transcripts, thirteen were known genes, 9 were predicted genes and all but one were located in the chromosome 4 QTL interval. Very interesting cis-regulated candidate genes for alcohol consumption were identified using microarray profiling of gene expression differences in congenic animals carrying a QTL for alcohol preference. Keywords: alcohol, congenic, rat, gene expression, brain, nucleus accumbens, amygdala, frontal cortex, hippocampus, striatum Overall design: Brain regions (accumbens, amygdala, frontal cortex, hippocampus, and striatum) from 6 biologic replicates of the selected non-preferring line NP and from a NP.P congenic line were dissected and subjected to microarray analysis for comparison.

在iP/iNP大鼠的4号染色体上，研究人员鉴定出一个影响酒精偏好的高度显著数量性状基因座（quantitative trait locus, QTL）。将iP大鼠4号染色体上的QTL区间导入iNP大鼠所构建的同源导入系（NP.P），与iNP背景品系相比，该品系表现出预期的酒精摄入量升高表型。本研究旨在鉴定4号染色体QTL区间内的基因，解析其在酒精未暴露的同源导入系与背景品系之间酒精摄入量差异中的潜在作用。研究人员从6只NP.P大鼠和6只iNP大鼠的5个脑区中提取RNA，分别进行标记后，在Affymetrix大鼠基因组230 2.0微阵列芯片（Affymetrix Rat Genome 230 2.0 microarray）上开展独立分析。采用稳健多芯片平均法（robust multi-chip average, RMA）对基因表达水平进行归一化处理，并通过实时定量聚合酶链式反应（quantitative real-time PCR）验证差异表达基因。对伏隔核（nucleus accumbens）、前额叶皮层（frontal cortex）、杏仁核（amygdala）、海马体（hippocampus）和纹状体（striatum）这5个脑区进行整合分析后，共鉴定出23个转录本与9个表达序列标签（expressed sequence tag, EST）在NP.P与iNP大鼠间存在差异表达。在这23个转录本中，13个为已知基因，9个为预测基因，除1个外其余均位于4号染色体QTL区间内。本研究通过对携带酒精偏好QTL的同源导入系动物的基因表达差异进行微阵列分析，鉴定出一批与酒精摄入量相关的极具研究价值的顺式调控候选基因。关键词：酒精偏好、同源导入系、大鼠、基因表达、脑组织、伏隔核、杏仁核、前额叶皮层、海马体、纹状体。实验设计：分别采集6只非偏好品系NP大鼠的生物学重复样本以及NP.P同源导入系大鼠的脑组织，解剖获取伏隔核、杏仁核、前额叶皮层、海马体和纹状体，随后开展微阵列分析以实现组间比较。