|
| Status |
Public on Mar 12, 2014 |
| Title |
26972c GmbHLHM1 rep1 |
| Sample type |
RNA |
| |
|
| Source name |
26972c GmbHLHM1
|
| Organism |
Saccharomyces cerevisiae |
| Characteristics |
strain/genotype: ammonium transport mutant (26972c)
transfection: pYES3 containing GmbHLHm1
|
| Treatment protocol |
12 hour exposure to 1 mM ammonium and 2% galactose
|
| Growth protocol |
Starter yeast cultures were initially prepared to ensure uniform cell growth. The 26972c yeast strain (mep1-1, mep2∆, Mep3, ura-) harboring GmbHLHm1-pYES3 or the pYES3 empty vector was inoculated in 20 ml of Grenson’s minimal yeast media supplemented with 2% w/v glucose and 0.1% (w/v) L-proline. The cultures were grown in 50 ml falcon tubes and incubated at 28°C and shaken at 200 rpm for 24 h. Ten replicate sterile 50 ml falcon tubes containing identical media were spiked with 1 ml of each yeast starter culture and incubated at 28°C at 200 rpm for 2 days. Log-phase cells were pelleted, washed in sterile Milli Q H2O twice and then resuspended in 20 ml of Grenson’s minimal yeast media supplemented with 2% (w/v) galactose and 0.5 mM (NH4)2SO4. The cells were incubated at 28°C and shaken at 200 rpm. The cell cultures were harvested after 12 h of incubation and washed once in 20 ml of ice-cold sterile Milli Q water.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using hot acidified phenol according to: Collart MA & Oliviero S (2001) Preparation of yeast RNA. Current protocols in molecular biology:13.12. 11-13.12. 15.
|
| Label |
Biotin
|
| Label protocol |
RNA labeling was performed by the Ramaciotti Centre for Gene Function Analysis (University of New South Wales, Sydney, Australia).
|
| |
|
| Hybridization protocol |
RNA hybridization to the Affymetrix GeneChip Yeast Genome 2.0 Arrays was performed by the Ramaciotti Centre for Gene Function Analysis (University of New South Wales, Sydney, Australia).
|
| Scan protocol |
Scanning was performed by the Ramaciotti Centre for Gene Function Analysis (University of New South Wales, Sydney, Australia.
|
| Data processing |
Raw CEL data files were imported into GeneSpring software and pre-processed using Robust Multichip Average (RMA) algorithm where the variation between the chips was normalized at the probe set level to obtain the ratio of expression to the median expression for each sample.
|
| |
|
| Submission date |
Mar 11, 2014 |
| Last update date |
Mar 12, 2014 |
| Contact name |
Brent N Kaiser |
| E-mail(s) |
[email protected]
|
| Phone |
+61883136650
|
| Organization name |
The University of Adelaide
|
| Department |
School of Agriculture Food and Wine
|
| Street address |
Waite Campus
|
| City |
Urrbrae |
| State/province |
SA |
| ZIP/Postal code |
5064 |
| Country |
Australia |
| |
|
| Platform ID |
GPL2529 |
| Series (1) |
| GSE55804 |
Expression data from 26972c yeast ± bHLHm1 (SAT1) |
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