Gene expression profiling of archival tongue carcinoma and normal tongue tissue (subset)

RNA extracted from 78 FFPE tongue samples, 62 tongue carcinomas and 16 non-malignant controls, were succesfully analysed using the whole genome DASL array to obtain gene expression profiles. Gene expression profiles were used to identify differentially expressed genes with the ultimate goal of finding out their importance for tongue cancer development and maintenance. Sample were formalin fixed paraffin embedded biopsies taken for diagnostic purposes and had been stored between 1 and 13 years. Because of the general poor quality of RNA from archival samples a special focus were put on its effect on the detected expression levels. 28 tumours and 16 controls with a CTdiff< 5 were selected for differential gene expression analysis. Raw data for only these samples were normalized again. Data for these 44 samples can be found here. Total RNA from formalin fixed paraffin embedded sample was extracted using the High Pure RNA Paraffin Kit according to the manufacturer’s protocol. Quality of FFPE sample were evaluated by comparing how well a kousekeeping gene (TUBA6) could be amplified using q-PCR in each FFPE sample compared to two fresh frozen sample (Ctdiff= CTffpe-Ctff). The DASL platform (cDNA-mediated annealing, selection, extension, and ligation assay) was used together with the whole genome arrays to obtain expression data for 20 818 genes.本研究针对78份舌部福尔马林固定石蜡包埋（FFPE）样本成功开展基因表达分析，其中包含62例舌癌组织样本与16例非恶性对照样本。实验采用全基因组DASL芯片完成检测，获取了所有样本的基因表达谱。研究人员基于所得基因表达谱鉴定差异表达基因，最终目标为明确这些基因在舌癌发生与维持进程中的关键作用。 本次研究所用样本均为用于诊断目的获取的福尔马林固定石蜡包埋活检组织，保存时长介于1至13年之间。鉴于存档样本提取的RNA普遍质量欠佳，本研究重点关注了RNA质量对检测所得表达水平的影响。最终筛选出28例肿瘤样本与16例对照样本（CTdiff<5）用于差异基因表达分析，并对这44份样本的原始数据再次进行标准化处理，相关数据可在此处获取。 福尔马林固定石蜡包埋样本的总RNA提取采用High Pure RNA Paraffin Kit，并严格遵循厂商提供的实验流程。FFPE样本的质量通过以下方式评估：以两份新鲜冷冻样本作为参照，采用q-PCR扩增管家基因TUBA6，计算CTdiff值（CTdiff= CTffpe - Ctff），以此评估各FFPE样本的RNA扩增效果与质量。 本研究采用DASL平台（即cDNA介导退火、选择、延伸与连接检测技术）结合全基因组芯片，可同时获取20818个基因的表达数据。