|
| Status |
Public on Sep 19, 2013 |
| Title |
M9-without nickel-log phase-rep1 |
| Sample type |
RNA |
| |
|
| Source name |
Nickel-resistant Saccharomyces cerevisiae, log phase
|
| Organism |
Saccharomyces cerevisiae |
| Characteristics |
phenotype: nickel-resistant mutant
|
| Treatment protocol |
Yeast cells were not treated with any special agent.
|
| Growth protocol |
Fresh precultures of yeast cells were diluted to an OD600 of 0.1 in fresh minimal medium. They were cultivated at 30°C and 150 rpm untill they reach the OD600 value of about 1.00. Cells were harvested at this OD600 value.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA of yeast cells were extracted by using RNeasy Mini Kit supplied from QIAGEN according to the manufacturer's recommendations. RNA was quantified using a NanoDrop-2000 spectrophotometer and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA).
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled RNA was prepared from 0.5 µg RNA using the One-color Low Input Quick Amp Kit (Agilent) according to the manufacturer's instructions, followed by Agilent Nano-prep RNA purification (Agilent, Santa Clara, CA). Dye incorporation and cRNA yield were checked with the NanoDrop-2000 spectrophotometer.
|
| |
|
| Hybridization protocol |
0.6 µg of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/µg cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 25 µl containing 25x Agilent fragmentation buffer and 10x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 25 µl of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Yeast V2 Oligo Microarrays (G4813A-016322) for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent).
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505B) using one color scan setting for 1x15k array slides (Scan Area 61x21.6 mm, Scan resolution 5µm, Dye channel is set to Green and Green PMT is set to 100%).
|
| Description |
Nickel-resistant Saccharomyces cerevisiae, log phase, replicate 1
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 9.1 (Agilent) using default parameters (protocol GE1-v1_91 and Grid: 016322_D_F_20070321). Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded.
|
| |
|
| Submission date |
Sep 18, 2013 |
| Last update date |
Sep 19, 2013 |
| Contact name |
CEREN ALKIM |
| E-mail(s) |
[email protected]
|
| Organization name |
INRA
|
| Street address |
Laboratoire d'Ingénierie des Systèmes Biologiques & des Procédés
|
| City |
TOULOUSE |
| ZIP/Postal code |
31077 |
| Country |
France |
| |
|
| Platform ID |
GPL16244 |
| Series (1) |
| GSE50985 |
Transcriptomic analysis of a nickel-resistant Saccharomyces cerevisiae mutant obtained by evolutionary engineering |
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