|
| Status |
Public on Mar 18, 2014 |
| Title |
Mutant+H2O2 |
| Sample type |
RNA |
| |
|
| Source name |
Yeast sod1Δ mutant with H2O2 treatment
|
| Organism |
Saccharomyces cerevisiae |
| Characteristics |
strain: FM391
ploidy: Haploid
agent: H2O2
|
| Treatment protocol |
Cells were treated with 0.4 mM H2O2 for 20 min.
|
| Growth protocol |
Cells were grown in yeast synthetic complete drop out (SC-Leu) medium at 30 °C on shaker to early log phase (OD600 ~0.4).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cells were resuspended in TES solution (10 mM Tris-HCl [pH 7.5], 10 mM EDTA, 0.5% SDS), and incubated with acidic phenol (pH 4.5; Sigma) for 1 hr at 65 °C. Samples were placed on ice for 10 min and centrifuged for 5 min at 13,000 rpm at 4 °C. Aqueous phases were extracted again with phenol and chloroform. RNA was precipitated in 100% ethanol for at least 2 h at -20 °C, centrifuged at 13,000 rpm for 15 min at 4°C, washed with 70% ethanol, and resuspended in water. Residual DNA was removed by DNase digestion using the RNase-Free DNase Set (Qiagen). RNA samples were further purified by RNeasy Kit (Qiagen).
|
| Label |
Biotin
|
| Label protocol |
RNA samples were amplified and reverse-transcribed using the NuGEN Ovation Pico WTA System V2 with 50ng RNA according to the manufacturer’s protocol. The yield of resulting cDNA was measured on the Caliper LabChip DS, and product sizing was evaluated on the Bioanalyzer. 5µg of each sample was fragmented and labeled with the Nugen Encore Biotin Module following the manufacturer’s protocol.
|
| |
|
| Hybridization protocol |
Samples were prepared for hybridization to the Affymetrix Yeast Genome 2.0 arrays using the Affymetrix GeneChip Hybridization, Wash and Stain Kit. Modifications were made to the Affymetrix protocol using the cocktail assembly for Mini Arrays. Samples were hybridized overnight in an Affymetrix GeneChip Hybridization Oven 640.
|
| Scan protocol |
Samples were then processed on the GeneChip Fluidics Station 450 and GeneChip Scanner 3000 7G.
|
| Description |
Gene expression data from early log phase of cells
|
| Data processing |
Data were analyzed with Genesifter software, Analysis Edition (Perkin Elmer) for background correction and GC-RMA was used as the normalization method.
|
| |
|
| Submission date |
Feb 17, 2014 |
| Last update date |
Mar 19, 2014 |
| Contact name |
Chi Kwan Tsang |
| E-mail(s) |
[email protected]
|
| Organization name |
Jinan University
|
| Department |
Clinical Neuroscience Institute
|
| Street address |
Building #3, 613 Huangpu avenue west
|
| City |
Guangzhou |
| ZIP/Postal code |
510630 |
| Country |
China |
| |
|
| Platform ID |
GPL2529 |
| Series (1) |
| GSE55081 |
Expression data from yeast wild type (WT) and sod1Δ strains after treatment with hydrogen peroxide (H2O2) |
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