应用基因芯片技术筛选胰腺癌相关基因

Screening the human pancreatic cancer associated genes by cDNA microarray

目的应用基因芯片技术筛选胰腺癌相关基因。方法将14000种人类基因PCR产物按微矩阵排列点样于化学涂层的载玻片上,制成基因芯片。按一步法抽提4例胰腺癌和癌旁正常胰腺组织的总RNA,将等量的RNA分别逆转录合成荧光分子掺入的cDNA一链作探针,混合后杂交上述基因芯片。经严格洗片后用ScanArray4000扫描仪扫描芯片荧光信号图像,每点上两种荧光信号的强度分别代表Cy5-dCTP和Cy3-dCTP的量,获得的荧光信号图像用计算机分析。结果按差异显著性标准,从14000个基因中筛选出在胰腺癌组织中共同差异表达基因189条,其中已知基因101条,新基因88条。在筛选出的已知基因中,有50条表达上调,51条表达下调。结论基因芯片技术的肿瘤基因表达谱分析能够高通量筛选胰腺癌相关基因,并高效对基因功能进行研究。胰腺癌基因表达谱的分析有助于认识肿瘤发病机制。

Objective To screen the human pancreatic cancer associated genes by cDNA microarray. Methods The PCR products of 14 000 genes were spotted onto a chemical-material-coated-glass plate in array. DNAs were fixed onto the glass plate after a series of treatments.Total RNAs were isolated from 4 pecimens of the pancreatic cancer and the normal tumor-surrounding pancreatic tissues. Both equal quantity RNAs were reversely transcribed to cDNAs and labeled with the fluorescent Cy5-dCTP and Cy3-dCTP to prepare the hybridization probes. The mixed probes were hybridized to the cDNA microarray. After high-stringent washing, the fluorescent signals were scanned by ScanArray 4000 scanner. The values of Cy5-dCTP and Cy3-dCTP on each spot were analyzed and calculated. Results By applying this cDNA microarray, 189 differentially expressed genes in pancreatic cancer, whose ratios of Cy5/Cy3 were higher than 2.0 or lower than 0.5, were screened out among the 14 000 target genes, comprising 101 known genes and 88 novel genes. Among the known genes, upregulated and downregulated genes were 50 and 51, respectively. Conclusions The analysis of gene expression profile of tumor based on cDNA microarray can realize high-throughput screening of the genes associated with the pancreatic cancer, and help to explore the gene function rapidly. Further analysis of the obtained genes will help to understand the molecular mechanism of the pancreatic cancer.