Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Normal vs. Tumor Prostate miRNA

ABSTRACT: Comparison of miRNA expression profiles in a small set of prostate needle core biopsies or fine needle aspirates. Keywords: Expression profiling of prostate needle core biopsies MicroRNA expression was compared between a pooled normal sample consisting of 10 separate normal adjacent to tumor prostate needle core biopsies, two prostate tumor cell lines (PC3 and LNCaP), two needle core biopsies, and a fine needle aspirate of a prostate tumor metastasis to the supraclavicular lymph node. MicroRNA was isolated from fresh frozen tissue sections of the needle core biopsies using the mirVana miRNA Isolation kit from Ambion per the manufacturer's instructions. MicroRNA was amplified using 10 ng input and 750 ng of amplified material was subsequently labeled for hybridization. All samples were normalized to the same normal prostate control.

ORGANISM(S): Homo sapiens

SUBMITTER: Michael Mattie

PROVIDER: E-GEOD-4574 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Optimized high-throughput microRNA expression profiling provides novel biomarker assessment of clinical prostate and breast cancer biopsies.

Mattie Michael D MD Benz Christopher C CC Bowers Jessica J Sensinger Kelly K Wong Linda L Scott Gary K GK Fedele Vita V Ginzinger David D Getts Robert R Haqq Chris C

Molecular cancer 20060619

<h4>Background</h4>Recent studies indicate that microRNAs (miRNAs) are mechanistically involved in the development of various human malignancies, suggesting that they represent a promising new class of cancer biomarkers. However, previously reported methods for measuring miRNA expression consume large amounts of tissue, prohibiting high-throughput miRNA profiling from typically small clinical samples such as excision or core needle biopsies of breast or prostate cancer. Here we describe a novel ...[more]

PMID: 16784538

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Project description:Omeprazole stimulated gene expression in rats Samples and RNA preparation: The series is composed of 9 hybridizations for analysis of differentially expressed genes in the gastric mucosa of omeprazole dosed vs. control rats. Nine rats (200-250 g) were given (by gavage) 400 micromol/kg of the proton pump inhibitor omeprazole (Astra AB, Gothenburg, Sweden) in Methocel (Dow Corning, Midland, MI) daily for 10 weeks. Seven rats received vehicle only. After 10 weeks the stomachs were removed for sampling of full-thickness corpus from the upper part of the greater curvature. Total RNA extraction was performed first by ultrasentrifugation on a cesium chloride cushion and then using TRIZOL Reagent (GIBCO BRL Life Technologies, New York, NY). Labeling and hybridization: Total RNA from 7 untreated rats were pooled and used as a common control. Three micrograms of total RNA from the control pool, and three micrograms of total RNA from each omeprazole dosed rat were reverse transcribed and labeled with capture sequences for subsequent attachment of Cy3- and Cy5-attached dendrimer, respectively, using 3DNA Array 350 Expression Array Detection kit as described in the manufacturer's protocols (Genisphere, Montvale, NJ). The labeling reactions were also spiked with exogenous control RNAs (Stratagene) at ratios of 1.0, 2.0, 3.0, 4.0, 5.0, 6.0, 0.5, 0.33, 0.25 and 0.125 for control spike 1, 2, 3, 4, 5, 6, 8, 9, 10 and 7, respectively. The slides were pre-washed in 2xSSC, 0.2% SDS at 55oC for 20 min, then 0.2x SSC at room temperature for 3 min, and deionized sterile filtered H2O at room temperature for 2 min. The Cy3 and Cy5 labeled samples (combined volume 9 microliter) were mixed with 25 microliter of the hybridization solution containing 0.25 M NaPO4, 1 mM EDTA, 4.5% SDS, 1 x SSC, 2 x Denhardt's solution and 2 microliter LNA dT Blocking reagent (Genisphere). A total hybridization volume of 50 microliter was added onto the array and covered with 24 x 60 mm cover slip. The array was assembled into humidified hybridization chamber (Corning) and hybridized at 60 oC for 18 h by submerging in a water bath. Post-hybridization washes were done once in 2 x SSC, 0.2% SDS at 55 oC for 15 min, then in 2 x SSC at room temperature for 15 min, and finally with 0.2 x SSC at room temperature for 15 min. After washing the fluorescent 3DNA reagent was hybridized to the cDNA at 60 oC for 3 h as described in the manufacturer's protocols. After dendrimer hybridization the slides were washed as described above. Scanning, image analysis and data processing: Arrays were scanned at 10 micrometer resolution with a ScanArray Express HT scanner (Packard BioSciences, Billerica, MA). The microarrays were analyzed using GenePixâ¢ Pro 4.1 (Axon Instruments, Inc., Union City, CA). All subsequent statistical analyses were performed using the statistical package R, R Development Core Team. A signal intensity based criterion was applied to remove the spots with median spot signal intensity less than 200. A spot area based criterion was applied to remove the spots with the area less than 50 pixels in either channel. Spots with more than 40% of the pixels saturated in either channel were also removed. Print tip-dependent normalization was applied to the data in order to compensate for systematic errors. The normalized ratios of the duplicated spots were averaged. Further analysis was done on the log2 transformed ratios. In order to assess the significance of up- or downregulation of the genes, tests for differential expression were performed using moderated T-tests, as implemented in the Limma R package of Smyth.

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Dataset Information

Normal vs. Tumor Prostate miRNA

Publications

Optimized high-throughput microRNA expression profiling provides novel biomarker assessment of clinical prostate and breast cancer biopsies.

Similar Datasets

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