Transcriptome profile of breast cancer HS-578T cells overexpressing EP300
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PROVIDER: PRJNA478310 | ENA |
REPOSITORIES: ENA
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Project description:EP300, a transcriptional co-activator of E-cadherin, has been recently found by our group to regulate doxorubicin resistance via by-pass of senescence and paclitaxel resistance by overcoming apoptosis in a minimally transformed mammary epithelial cells (MTMEC). Moreover, EP300 deleted MTMEC cells exhibit an multi-drug resistant (MDR) phenotype independent of P-glycoprotein (ABCB1), an efflux pump or ABC drug transporter. This whole transcriptome array study was undertaken in order to explore the downstream targets in the EP300-mediated drug resistance, epidermal-to-mesenchymal transition and cancer stem cell phenotypes in breast cancer cell line MCF7.
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Project description:Gene expression of different flocculent (HS 10 and HS 34) and powdery (HS 12 and HS 37) yeast strains compared to each other during exponential and stationary growth phase was analysed. The isolation of RNA was done by disruption of the cells under liquid nitrogen using mortar and pistil and then the Qiagen RNeasy Midi Kit with some modifications within the manufacturers´ protocol and the Qiagen RNase-Free DNase Set were applied. 6 µg of the total RNA per sample was used for each microarray experiments. The indirect labelling by the tyramide-signal-amplification method (MicromaxTM TSATM labelling and detection Kit from Perkin Elmer life sciences) was used to increase the Cy3 and Cy5 signals of microarray detection. Each cDNA containing Biotin- and Fluorescein-nucleotides respectively was purified with a QIAquick PCR purification kit and suspended in 11 µl of the formamide containing hybridization buffer. The slides were hybridized at 42°C over night under a cover slip. The microarrays were scanned by the Axon 4000B scanner; image intensity data were extracted and analysed with GenePix® Pro 6.0 software. Data from different scans of Dye-swap experiment were extracted by GenePix Pro 6.0 software, normalized and united. An outliertest has been applied in order to find outliers amongst the gene replicats. Subsequently, a t-test (1% and 5% probability of error) has been used in order to find regulated genes. Keywords: sorted yeast cells Microarray experiments were realised with dye swap.
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Project description:Gene expression of different flocculent (HS 10 and HS 34) and powdery (HS 12 and HS 37) yeast strains compared to each other during exponential and stationary growth phase was analysed. The isolation of RNA was done by disruption of the cells under liquid nitrogen using mortar and pistil and then the Qiagen RNeasy Midi Kit with some modifications within the manufacturers´ protocol and the Qiagen RNase-Free DNase Set were applied. 6 µg of the total RNA per sample was used for each microarray experiments. The indirect labelling by the tyramide-signal-amplification method (MicromaxTM TSATM labelling and detection Kit from Perkin Elmer life sciences) was used to increase the Cy3 and Cy5 signals of microarray detection. Each cDNA containing Biotin- and Fluorescein-nucleotides respectively was purified with a QIAquick PCR purification kit and suspended in 11 µl of the formamide containing hybridization buffer. The slides were hybridized at 42°C over night under a cover slip. The microarrays were scanned by the Axon 4000B scanner; image intensity data were extracted and analysed with GenePix® Pro 6.0 software. Data from different scans of Dye-swap experiment were extracted by GenePix Pro 6.0 software, normalized and united. An outliertest has been applied in order to find outliers amongst the gene replicats. Subsequently, a t-test (1% and 5% probability of error) has been used in order to find regulated genes. Keywords: sorted yeast cells
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