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Real-time quantitative PCR analysis of human colonic HT-29 cells under insulin stimulation

ABSTRACT: Compared mRNA expression between control and insulin treated cells. HT29 cells were treated with 100nM Insulin for 24h. The RT2 Profiler PCR Arrays PAHS-30C (SA Biosciences, MD, USA) was designed to analyze 84 genes related to human insulin signaling pathway. The RT-PCR was carried out using an ABI PRISM® 7000 Sequence Detection System (Applied Biosystems, Foster City, CA).The expression of HSD11B2 for the 3 experiments concerned was monitored in parallel by real time PCR which confirming significant downregulation by insulin.

ORGANISM(S): Homo sapiens

PROVIDER: GSE51677 | GEO | 2014/06/01

SECONDARY ACCESSION(S): PRJNA224313

REPOSITORIES: GEO

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Real-time quantitative PCR analysis of human colonic HT-29 cells under insulin stimulation

Project description:Compared mRNA expression between control and insulin treated cells. HT29 cells were treated with 100nM Insulin for 24h. The RT2 Profiler PCR Arrays PAHS-30C (SA Biosciences, MD, USA) was designed to analyze 84 genes related to human insulin signaling pathway. The RT-PCR was carried out using an ABI PRISMM-BM-. 7000 Sequence Detection System (Applied Biosystems, Foster City, CA).The expression of HSD11B2 for the 3 experiments concerned was monitored in parallel by real time PCR which confirming significant downregulation by insulin. qPCR gene expression profiling. RNA from 3 separated experiments was collected.

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Real-time quantitative PCR analysis of 88 microRNAs involved in human cell differentiation and development in systemic sclerosis (SSc) dermal fibroblasts

Project description:Normal fibroblasts and SSc fibroblasts between the third and six subpassages were used for experiments. Total RNA was extracted from culture cells with ISOGEN (Nippon Gene, Tokyo, Japan). MicroRNA isolation from total RNA was performed using RT2 qPCR-Grade miRNA Isolation Kit (SA Bioscience). For RT2 Profiler PCR Array (SABioscience), microRNAs were reverse-transcribed into first strand cDNA using RT2 miRNA First Strand Kit (SABiosciences). A mixture of equal amounts of cDNAs from 5 normal fibroblasts or 5 SSc fibroblasts was prepared. The cDNA was mixed with RT2 SYBR Green/ROX qPCR Master Mix and the mixture was added into a 96-well RT2 miRNA PCR Array (SABiosciences) that included primer pairs for 88 human microRNAs.

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Project description:Human osteoarthritic (OA) chondrocytes were obtained from the cartilage of femoral condyles and tibial plateaus from three patients undergoing total knee arthroplasty. The chondrocytes were released from the cartilage, seeded at high density, and cultured in Dulbecco’s modified Eagle’s medium supplemented with 10% heat-inactivated fetal bovine serum and an antibiotic mixture (100 units/ml penicillin base and 100 μg/ml streptomycin base) at 37°C in a humidified atmosphere of 5% CO2/95% air. The chondrocytes from each of the three donors were left untreated (control) and treated with tunicamycin (500ng/ml for 20 hours).Total RNA was extracted from the chondrocytes and processed for qPCR according to the manufacturer's instructions (SA Biosciences RT2 Profiler PCR Array PAHS-089Z).

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