|
| Status |
Public on Jan 10, 2017 |
| Title |
rapamycine H2O vs rapamycine NH4 replicate 2 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Total RNA of S288C strain
|
| Organism |
Saccharomyces cerevisiae S288C |
| Characteristics |
addition at t0: H2O
biological rep: 2
|
| Growth protocol |
An overnight cell pre-culture of S288c yeast strain was firstly obtained on a YEPD medium, before to transfer cells in an YNB medium for 32h. A constant population of cells was then nitrogen starved for 16h in an YNB-N medium in order to reach the stationary phase. Ammonium chloride (20 mM) or water (control) was then added to the medium after supplementation with rapamycin.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
After 30 minNH4 or H2O addition, sampling was performed in less than 1 min as followed: cells were filtrated under vacuum on a 0.45 µm HA membrane, collected and suspended in 1.5 ml DEPC water then stored at -80°C before RNA extraction. RNA were extracted from yeast cell culture as described previously (Chomczynski and Sacchi, 1987). The RNA concentrations were determined by measuring absorbance at 260 nm with a Nanodrop apparatus.
|
| Label |
cy3
|
| Label protocol |
Retrotranscription and labeling were performed using a ChipShot direct labeling and clean-up system kit (Promega) according to manufacturer’s indications.
|
| |
|
| Channel 2 |
| Source name |
Total RNA of S288C strain
|
| Organism |
Saccharomyces cerevisiae S288C |
| Characteristics |
addition at t0: NH4
biological rep: 2
|
| Growth protocol |
An overnight cell pre-culture of S288c yeast strain was firstly obtained on a YEPD medium, before to transfer cells in an YNB medium for 32h. A constant population of cells was then nitrogen starved for 16h in an YNB-N medium in order to reach the stationary phase. Ammonium chloride (20 mM) or water (control) was then added to the medium after supplementation with rapamycin.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
After 30 minNH4 or H2O addition, sampling was performed in less than 1 min as followed: cells were filtrated under vacuum on a 0.45 µm HA membrane, collected and suspended in 1.5 ml DEPC water then stored at -80°C before RNA extraction. RNA were extracted from yeast cell culture as described previously (Chomczynski and Sacchi, 1987). The RNA concentrations were determined by measuring absorbance at 260 nm with a Nanodrop apparatus.
|
| Label |
cy5
|
| Label protocol |
Retrotranscription and labeling were performed using a ChipShot direct labeling and clean-up system kit (Promega) according to manufacturer’s indications.
|
| |
|
| |
| Hybridization protocol |
Microarray hybridizations were performed using a Pronto Universal Microarray kit (Corning) according to manufacturer’s indications. Two biological replicates were done based on a dye swap design (S288C strain, 2 biological replicates with ammonium chloride addition and 2 biological replicates with water addition (control)).
|
| Scan protocol |
Microarrays were scanned using a GenePix pro 3 scanner (Axon Instruments).
|
| Description |
Biological replicate 2 of 2 of rapamycine and NH4 addition vs. Biological replicate 2 of 2 of rapamycine and H2O addition. Swap 2
|
| Data processing |
Data were processed using the R software (R2.9.2) and the Limma package (R Development Core Team 2010; Smyth 2005, Smyth et al. 2005; Smyth and Speed 2003). Within and between array normalizations were performed respectively by the print-tip-loess and quantile methods. Differentially expressed genes were identified through a linear model approach and a Benjamini-Hochberg method was used to adjust the p-values (Benjamini and Hochberg, 1995).
|
| |
|
| Submission date |
Jan 11, 2016 |
| Last update date |
Jan 11, 2017 |
| Contact name |
Bruno Blondin |
| E-mail(s) |
bruno.blondin@supagro.inra.fr
|
| Organization name |
INRA
|
| Department |
UMR1083
|
| Lab |
SPO
|
| Street address |
2 place Pierre Viala
|
| City |
Montpellier |
| ZIP/Postal code |
34060 |
| Country |
France |
| |
|
| Platform ID |
GPL21316 |
| Series (2) |
| GSE76716 |
Global transcriptome changes after nitrogen replenishment following a rapamycin treatment in S288c yeast strain |
| GSE76778 |
Global transcriptome changes after nitrogen replenishment following or not a rapamycin treatment in S288c yeast strain |
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