基因表达谱芯片技术筛选涎腺腺样囊性癌ACC-2/BLM细胞耐药相关基因

Screening and identification of multidrug resistance-associated genes in salivary gland adenoid cystic carcinoma ACC-2/BLM cells by cDNA microarry

目的:应用高通量基因芯片技术对涎腺腺样囊性癌细胞系ACC-2及其多药耐药细胞株ACC-2/BLM的基因表达谱进行对比分析,筛选差异表达基因。方法:以涎腺腺样囊性癌细胞系ACC-2及其耐药细胞ACC-2/BLM作为研究对象,提取mRNA,逆转录为cDNA,用Cy3和Cy5荧光染料标记,制备成cDNA探针,与表达谱芯片进行杂交扫描和分析;并应用Real-Time-PCR技术对其中6个基因进行验证。结果:在45 015个基因表达谱的筛选中,2 294个基因表达差异(2倍以上),其中1 310个基因表达水平上调,984个基因表达水平下调。使用RT-PCR和RealTime PCR技术对其中6个基因表达差异进行验证,验证结果与基因芯片结果一致。结论:涎腺腺样囊性癌ACC-2/BLM细胞耐药性与多个基因相关。

Objective：To identify multidrug resistance-associated genes in salivary gland adenoid cystic carcinoma ACC-2/BLM cells.Methods：Salivary gland adenoid cystic carcinoma cell line ACC-2 and its multidrug resistant cell line ACC-2/BLM cells were used to screen the multidrug-resistance related genes in ACC by microarray technology.Two fluorescent cDNA probes labeled with Cy3 and Cy5 dyes were prepared from the mRNA samples of ACC-2 and ACC-2/BLM cells by reverse transcription method.The two color probes were then mixed and hybridized on the cDNA chip constructed by double dots of 45 015 human genes,and scanned at two wave lengths.Genes expressed at different levels of the two cell lines were analyzed using computer.Then six of the differently expressed genes were further validated by RT-PCR.Results：Of the 45 015 known genes and expressed sequence tags,2 292 genes showed significantly different expression level（minimum 2 fold） between the two cell lines.Among the 2 294 genes,1 310 were up regulated（with ratio more than 2） and 984 were down（with ratio less than 1/2）.The results of RT-PCR analysis for 6 differently expressed genes were coincident with those of microarray assay.Conclusion：The multidrug-resistance of ACC-2/BLM cells is associated with the expression of multiple genes.