|
| Status |
Public on Oct 10, 2012 |
| Title |
Wt threonine24h x alcAypkA threonine24h Replicate 2 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
total RNA from Wt grown on glycerol 2% threonine 100mM, labeled with Cyanine-3
|
| Organism |
Aspergillus nidulans |
| Characteristics |
genotype/variation: Wt
protocol: glycerol 2% threonine 100mM during 24h
|
| Growth protocol |
A. nidulans strains (Wt and alcAypkA) were grown on minimal medium with glucose 4% or glycerol 2% threonine 100mM during 16 or 24 hours at 37°C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using Trizol following manufacturer's instructions, followed by DNAse treatment and purification using RNeasy® Mini Kit (Qiagen).
|
| Label |
Cy3
|
| Label protocol |
Prior to labelling, cDNA was synthesized from 5 µg of RNA using Agilent's cDNA Master Mix. cRNA amplification and labeling were performed by adding to the samples the Agilent™ Transcription Master Mix plus Cyanine-3 (for untreated samples) or Cyanine-5 (for SEB-treated samples) for 2 hours at 40°C.
|
| |
|
| Channel 2 |
| Source name |
total RNA from alcAypkA grown on glycerol 2% threonine 100mM, labeled with Cyanine-5
|
| Organism |
Aspergillus nidulans |
| Characteristics |
genotype/variation: alcAypkA
protocol: glycerol 2% threonine 100mM during 24h
|
| Growth protocol |
A. nidulans strains (Wt and alcAypkA) were grown on minimal medium with glucose 4% or glycerol 2% threonine 100mM during 16 or 24 hours at 37°C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using Trizol following manufacturer's instructions, followed by DNAse treatment and purification using RNeasy® Mini Kit (Qiagen).
|
| Label |
Cy5
|
| Label protocol |
Prior to labelling, cDNA was synthesized from 5 µg of RNA using Agilent's cDNA Master Mix. cRNA amplification and labeling were performed by adding to the samples the Agilent™ Transcription Master Mix plus Cyanine-3 (for untreated samples) or Cyanine-5 (for SEB-treated samples) for 2 hours at 40°C.
|
| |
|
| |
| Hybridization protocol |
For the hybridization, 825 ng of each labeled cRNA was mixed with Agilent™ Fragmentation Mix and incubated at 60 °C for exactly 30 minutes to fragment RNA. The fragmentation was interrupted by adding 55 µL of 2X GE Hybridization Buffer HI-RPM. Finally, 100 µL of sample was placed down onto the microarray slide, which was mounted into the Agilent™ Microarray Hybridization Chamber Kit. The hybridization was carried out in an oven (Agilent G2545A Hybridization Oven) set to 65 °C for 17 hours. After, microarray slides were washed according to Agilent’s instruction
|
| Scan protocol |
Scanned using GenePix® 4000B microarray scanner (Molecular Devices, USA).
|
| Description |
Biological replicate 2 of 2.
|
| Data processing |
Agilent Feature Extraction Software (v 9.5.3.1) was used for background subtraction and LOWESS normalization. After data were processed using TIGR platform. Data were visualized using TMeV from TIGR, and the same software has been used for statistical analysis (t test).
|
| |
|
| Submission date |
Jul 30, 2012 |
| Last update date |
Oct 10, 2012 |
| Contact name |
Ana Cristina Colabardini |
| E-mail(s) |
anacolabardini@gmail.com
|
| Phone |
551636024311
|
| Organization name |
FCFRP-USP
|
| Department |
Ciências Farmacêuticas
|
| Lab |
Laboratório de Bilogia Molecular
|
| Street address |
Av do Café s/n Cidade Universitária
|
| City |
Ribeirão Preto |
| State/province |
São Paulo |
| ZIP/Postal code |
14025110 |
| Country |
Brazil |
| |
|
| Platform ID |
GPL15870 |
| Series (1) |
| GSE39755 |
Aspergillus nidulans: Control (Wt) vs. ypkA induction or repression (alcAypkA) |
|
