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

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. Overall design: 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.

本研究从78份舌部福尔马林固定石蜡包埋（FFPE）样本中提取RNA，其中包含62份舌癌组织样本与16份非恶性对照样本，成功采用全基因组DASL芯片完成分析，获取基因表达谱。基于所得基因表达谱鉴定差异表达基因，最终旨在阐明其在舌癌发生与维持进程中的重要作用。所用样本为用于临床诊断的福尔马林固定石蜡包埋活检组织，存储时长介于1至13年不等。鉴于存档样本的RNA普遍质量较差，本研究重点关注其对检测所得表达水平的影响。最终筛选出28份肿瘤样本与16份对照样本（CTdiff<5）用于差异基因表达分析，并对这44份样本的原始数据重新进行标准化处理。上述44份样本的相关数据可于此处获取。实验整体设计如下：按照制造商提供的操作手册，使用High Pure RNA Paraffin Kit提取福尔马林固定石蜡包埋样本中的总RNA；通过比较每份FFPE样本与两份新鲜冷冻样本中管家基因TUBA6的实时定量PCR（q-PCR）扩增效果，评估FFPE样本的RNA质量，其中Ctdiff= CTffpe - CTff；采用搭载全基因组芯片的DASL技术平台（即cDNA介导退火、选择、延伸与连接检测技术），获取20818个基因的表达数据。