Project description:PURPOSE: To provide a detailed gene expression profile of the normal postnatal mouse cornea. METHODS: Serial analysis of gene expression (SAGE) was performed on postnatal day (PN)9 and adult mouse (6 week) total corneas. The expression of selected genes was analyzed by in situ hybridization. RESULTS: A total of 64,272 PN9 and 62,206 adult tags were sequenced. Mouse corneal transcriptomes are composed of at least 19,544 and 18,509 unique mRNAs, respectively. One third of the unique tags were expressed at both stages, whereas a third was identified exclusively in PN9 or adult corneas. Three hundred thirty-four PN9 and 339 adult tags were enriched more than fivefold over other published nonocular libraries. Abundant transcripts were associated with metabolic functions, redox activities, and barrier integrity. Three members of the Ly-6/uPAR family whose functions are unknown in the cornea constitute more than 1% of the total mRNA. Aquaporin 5, epithelial membrane protein and glutathione-S-transferase (GST) omega-1, and GST alpha-4 mRNAs were preferentially expressed in distinct corneal epithelial layers, providing new markers for stratification. More than 200 tags were differentially expressed, of which 25 mediate transcription. CONCLUSIONS: In addition to providing a detailed profile of expressed genes in the PN9 and mature mouse cornea, the present SAGE data demonstrate dynamic changes in gene expression after eye opening and provide new probes for exploring corneal epithelial cell stratification, development, and function and for exploring the intricate relationship between programmed and environmentally induced gene expression in the cornea. Keywords: other
Project description:In this study we have performed expression analysis using paired FF-FFPE glioma samples. We show that expression data from FFPE glioma material is concordant with expression data from matched FF tissue, and can be used for molecular profiling in gliomas. In this study we have performed expression analysis using 55 paired FF-FFPE glioma samples (HU133 plus 2.0 arrays (FF) and Human Exon 1.0 ST arrays (FFPE)). The most informative probe sets were selected based on variance This Series contains the FFPE data only. FF data set was previously submitted (GSE16011).
Project description:This study was performed to evaluated RNA extraction and gene expression analysis of FFPE specimen stored for more than 20 years. Using long time stored FFPE material; large retrospective studies correlating molecular features with therapeutic response and clinical outcome, can be performed. Quantitative PCR (qPCR) was used to evaluate RNA extraction methods and to compare gene expression profiles of FFPE and fresh frozen (FF) tissue. Extracted RNA was subsequently subjected to microarray analysis and compared to qPCR data. The Ambion RecoverAll kit appears to be particularly suited for RNA extraction of long time stored FFPE tissues. Gene expression analysis using Affymetrix platform displayed a high degree of correlation for endogenous control genes comparing FF and FFPE tissues. We conclude that high quality gene expression signatures can be recognized using Affymetrix gene expression platform on FFPE tissue stored for more than 20 years. However, a general interpretation must be done with caution as different FFPE procedures have varying effects on RNA quality. Three RNA extraction methods from Roche (Basel, Switzerland), Ambion (Austin, TX, USA) and Qiagen (Hilden, Germany), designed for FFPE material was used. The methods were compared using qPCR. For the qPCR analysis, two different concentrations of input cDNA were used (20ng and 100ng) and 32 human endogenous control genes were examined. RNA from the Ambion FFPE kit was further analyzed by using microarray. Amplification of RNA prior to microarray analysis was performed using Nugen technologies (San Carlos, CA, USA). Nugen has developed amplification kits both for FFPE and FF materials: WT- ovation FFPE RNA amplification System V2 (Nugen FFPE), Ovation FF RNA Amplification System V2 (Nugen V2 FF) and Ovation FF Pico RNA Amplification System (Nugen PICO FF). The Nugen FFPE kit, designed for FFPE material was only used for FFPE tissue. The Nugen Pico FF kit, designed to target small amounts of FF RNA (>500 pg) and the Nugen V2 FF kit designed to target total FF RNA, were only used for FF tissue. Affymetrix standard amplification protocol (Affy FF) designed for FF RNA was also included as the standard method for amplification.
Project description:In this study we have performed expression analysis using paired FF-FFPE glioma samples. We show that expression data from FFPE glioma material is concordant with expression data from matched FF tissue, and can be used for molecular profiling in gliomas.
Project description:This experiment contains the RNA-Seq samples only. Formalin-fixed, paraffin-embedded (FFPE) tissues are an invaluable resource for clinical research. However, nucleic acids extracted from FFPE tissues are fragmented and chemically modified making them challenging to use in molecular studies. We analysed 23 fresh-frozen (FF), 35 FFPE and 38 paired FF/FFPE specimens, representing six different human tissue types (bladder, prostate and colon carcinoma; liver and colon normal tissue; reactive tonsil) in order to examine the potential use of FFPE samples in next-generation sequencing (NGS) based retrospective and prospective clinical studies. Two methods for DNA and three methods for RNA extraction from FFPE tissues were compared and were found to affect nucleic acid quantity and quality. DNA and RNA from selected FFPE and paired FF/FFPE specimens were used for exome and transcriptome analysis. Preparations of DNA Exome-Seq libraries was more challenging (29.5 % success) than that of RNA-Seq libraries, presumably because of modifications to FFPE tissue-derived DNA. Libraries could still be prepared from RNA isolated from two-decade old FFPE tissues. Data were analysed using the CLC Bio Genomics Workbench and revealed systematic differences between FF and FFPE tissue-derived nucleic acid libraries. In spite of this, pairwise analysis of DNA Exome-Seq data showed concordance for 70-80 % of variants in FF and FFPE samples stored for fewer than three years. RNA-Seq data showed high correlation of expression profiles in FF/FFPE pairs (Pearson Correlations of 0.90 +/- 0.05), irrespective of storage time (up to 244 months) and tissue type. A common set of 1,494 genes was identified with expression profiles that were significantly different between paired FF and FFPE samples irrespective of tissue type. Our results are promising and suggest that NGS can be used to study FFPE specimens in both prospective and retrospective archive-based studies in which FF specimens are not available.
Project description:Comparison of gene expression profiles from Mus musculus brain at age 30 months. The RNA-seq data comprise 1 groups. Jena Centre for Systems Biology of Ageing - JenAge (www.jenage.de)
Project description:Comparison of gene expression profiles from Mus musculus brain (hemisphere) of animals kept in standard environment and enriched environment. The RNA-seq data comprise 4 groups: 2 age groups, each w/ and w/o enriched environment. Jena Centre for Systems Biology of Ageing - JenAge (www.jenage.de)
Project description:Comparison of gene expression profiles from Mus musculus brain (hippocampus) of animals kept in standard environment and enriched environment. The RNA-seq data comprise 4 groups: 2 age groups, each w/ and w/o enriched environment. Jena Centre for Systems Biology of Ageing - JenAge (www.jenage.de)