|
| Status |
Public on Feb 10, 2016 |
| Title |
Crz1 Overexpression pREP1 Replicate 2 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
empty vector pREP1 auxotrophic strain, untreated
|
| Organism |
Schizosaccharomyces pombe |
| Characteristics |
strain/background: JK366 (ade6-M216, leu1-32, ura4-D18) h-
genotype/variation: empty vector pREP1
treatment: none
|
| Growth protocol |
S. pombe yeast cells were inoculated in EMM+AU minimal media with no thiamine and cultured for 18-22hr.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using hot phenol-chloroform method, followed by mRNA isolation using gravity-flow Oligo-dT cellulose beads.
|
| Label |
Cy5
|
| Label protocol |
2 ug of mRNA were converted into cDNA using 140 nmol of Oligo-dT(25) anchored primer (Sigma), 20 nmol of aminoallyl dUTP dNTP (Sigma) mixture, and 200 units of SuperScript II reverse transcriptase (Invitrogen). 1 ug of cDNA each from overexpression or deletion strain and empty vector control strain was labeled with Cy3 or Cy5 (GE Healthcare) and were combined for QIAquick purification (QIAGEN).
|
| |
|
| Channel 2 |
| Source name |
Crz1 OE pREP1 auxotrophic strain, untreated
|
| Organism |
Schizosaccharomyces pombe |
| Characteristics |
strain/background: JK366 (ade6-M216, leu1-32, ura4-D18)
genotype/variation: Crz1 OE pREP1
treatment: none
|
| Growth protocol |
S. pombe yeast cells were inoculated in EMM+AU minimal media with no thiamine and cultured for 18-22hr.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using hot phenol-chloroform method, followed by mRNA isolation using gravity-flow Oligo-dT cellulose beads.
|
| Label |
Cy3
|
| Label protocol |
2 ug of mRNA were converted into cDNA using 140 nmol of Oligo-dT(25) anchored primer (Sigma), 20 nmol of aminoallyl dUTP dNTP (Sigma) mixture, and 200 units of SuperScript II reverse transcriptase (Invitrogen). 1 ug of cDNA each from overexpression or deletion strain and empty vector control strain was labeled with Cy3 or Cy5 (GE Healthcare) and were combined for QIAquick purification (QIAGEN).
|
| |
|
| |
| Hybridization protocol |
Hybridization was performed according to the manufacturer's instruction (Agilent Technology). Microarray was washed in 6X SSPE/0.005% sodium N-lauroylsarcosine at room temperature for 5 min, followed by a second wash in pre-heated 42°C 0.6X SSPE for 2 min.
|
| Scan protocol |
Microarrays were scanned on a GenePix 4200A instrument with GenePix 3.0 software.
|
| Description |
CRZ1 OE pREP1-rep2
Untreated strains.
|
| Data processing |
The raw data was lowess normalized and the average log2 ratios with the corresponding t-test p values from the dye-swap experiments were obtained using the R Bioconductor Limma package.
|
| |
|
| Submission date |
Feb 09, 2016 |
| Last update date |
Feb 10, 2016 |
| Contact name |
Gordon Chua |
| E-mail(s) |
gchua@ucalgary.ca
|
| Phone |
403-220-7769
|
| Organization name |
University of Calgary
|
| Department |
Biological Sciences
|
| Lab |
Institute for Biocomplexity and Informatics
|
| Street address |
2500 University Drive NW
|
| City |
Calgary |
| State/province |
Alberta |
| ZIP/Postal code |
T2N 1N4 |
| Country |
Canada |
| |
|
| Platform ID |
GPL16187 |
| Series (2) |
| GSE77759 |
Genome-wide analysis of Prz1 in fission yeast reveals a novel inhibitory role in flocculation and a conserved activating role in cell wall organization [expression] |
| GSE77761 |
Conserved and diverged functions of the calcineurin-activated Prz1 transcription factor in fission yeast |
|
