|
| Status |
Public on Apr 10, 2007 |
| Title |
2 Ol5h Rice Osmotic(Ol5h) Leaves 5 hours |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Osmotic(Ol5h)_Leaves_5_hours
|
| Organism |
Oryza sativa Japonica Group |
| Characteristics |
Tissue: 2-weeks old rice seedling Leaves. Treatment: Osmotic
|
| Biomaterial provider |
NA
|
| Treatment protocol |
100 mM of mannitol was added in the hydroponic solution. Root and shoot tissues were harvested 1 hr or 5 hr after the beginning of stress treatment
|
| Growth protocol |
Seeds of rice genotype Nipponbarre were obtained from the International Rice Research Institute (IRRI). Dehisced seeds were sterilized with 15% (v/v) sodium hypochlorite for 30 min and then rinsed with distilled water. Seeds were germinated in Petri dishes at 28oC on Whatman® paper soaked in deionized water. After 4 days, rice seedlings were transferred in pots containing pouzzolane and allowed to growth under controlled conditions with a 28°C/25°C day/night temperatures, 12 hr photoperiod and 55-65% humidity for 2 weeks in Yoshida’s modified solution (1981): 0.7 mM KNO3, 1.2 mM Ca(NO3)2, 1.6 mM MgSO4, 0.5 mM (NH4)2SO4, 0.8 mM KH2PO4, 60 µM FeEDTA, 20µM MnSO4, 0.32 µM(NH4)6Mo7O24, 1.4 µM ZnSO4, 1.6 µM CuSO4, 45.2 µM H3BO3. Medium pH was adjusted to 5.0 twice a day. Seedlings at the 5 leaves stage without any visible tiller were carefully selected for stress treatments.
Seeds of rice genotype Nipponbarre were obtained from the International Rice Research Institute (IRRI). Dehisced seeds were sterilized with 15% (v/v) sodium hypochlorite for 30 min and then rinsed with distilled water. Seeds were germinated in Petri dishes at 28oC on Whatman® paper soaked in deionized water. After 4 days, rice seedlings were transferred in pots containing pouzzolane and allowed to growth under controlled conditions with a 28°C/25°C day/night temperatures, 12 hr photoperiod and 55-65% humidity for 2 weeks in Yoshida’s modified solution (1981): 0.7 mM KNO3, 1.2 mM Ca(NO3)2, 1.6 mM MgSO4, 0.5 mM (NH4)2SO4, 0.8 mM KH2PO4, 60 µM FeEDTA, 20µM MnSO4, 0.32 µM(NH4)6Mo7O24, 1.4 µM ZnSO4, 1.6 µM CuSO4, 45.2 µM H3BO3. Medium pH was adjusted to 5.0 twice a day. Seedlings at the 5 leaves stage without any visible tiller were carefully selected for stress treatments.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
TRIZOL protocol (Invitrogen, Carlsbad, CA), as indicated by the manufacturer.
|
| Label |
Cy5
|
| Label protocol |
10 µg of total RNAs were used for Cy3 or Cy5 labelling using the Cyscribe First-Strand cDNA Labelling kit (Amersham). The labelled cDNA samples were purified using the CyScribe GFX Purification Kit (Amersham) and concentrated using a microcon YM-30 filter (Millipore). 5 ng of Luciferase RNA (Promega) was added to each RNA samples prior labelling and used as spiking / normalization control.!Sample_label_protocol_ch2 = 10 µg of total RNAs were used for Cy3 or Cy5 labelling using the Cyscribe First-Strand cDNA Labelling kit (Amersham). The labelled cDNA samples were purified using the CyScribe GFX Purification Kit (Amersham) and concentrated using a microcon YM-30 filter (Millipore). 5 ng of Luciferase RNA (Promega) was added to each RNA samples prior labelling and used as spiking / normalization control.
|
| |
|
| Channel 2 |
| Source name |
2_Leaves_5_hours_Mock_treated
|
| Organism |
Oryza sativa Japonica Group |
| Characteristics |
Tissue: 2-weeks old rice seedling Leaves. Treatment: Mock treatment
|
| Biomaterial provider |
NA
|
| Treatment protocol |
Water was added in the hydroponic solution. Root and shoot tissues were harvested 1 hr or 5 hr after the beginning of stress treatment
|
| Extracted molecule |
total RNA |
| Extraction protocol |
TRIZOL protocol (Invitrogen, Carlsbad, CA), as indicated by the manufacturer.
|
| Label |
Cy3
|
| |
|
| |
| Hybridization protocol |
For array hybridisation, Cy3 and Cy5 labelled cDNA probes were mixed with Calf thymus DNA (Sigma) and EGT hybridization buffer (Eurogentec) and hybridised to the microarray (Eurogentec) at 42°c in a humid chamber (Corning) for 16 hours. Arrays were washed 5min in a 0.2xSSC-0.1% sodium dodecyl sulfate solution then 5min in 0.2xSSC. The arrays were spin-dried.
|
| Scan protocol |
The arrays were scanned using the Axon 4100A Scanner. The hybridisation data were collected and normalization performed using the GenePix Pro 3.0 software.
|
| Description |
NA
|
| Data processing |
Data were normalized using Limma, a package of Bioconductor, using loess, but giving different weight do spots. Weight of each spot is reported in the data table.
|
| |
|
| Submission date |
Sep 13, 2006 |
| Last update date |
Feb 28, 2007 |
| Contact name |
Stefano Berri |
| Organization name |
University of Milan
|
| Department |
biomolecular sciences and biotechnology
|
| Street address |
via Celoria 26
|
| City |
Milan |
| ZIP/Postal code |
20133 |
| Country |
Italy |
| |
|
| Platform ID |
GPL4315 |
| Series (1) |
| GSE5819 |
Whole WRKY gene family transcription analysis in rice during biotic and abiotic stress. |
|
