|
| Status |
Public on Apr 10, 2007 |
| Title |
11 Or5h Rice Osmotic(Or5h) Roots 5 hours |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
11_Roots_5_hours_Mock_treated
|
| Organism |
Oryza sativa Japonica Group |
| Characteristics |
Tissue: 2-weeks old rice seedling Roots. 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
|
| 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.
|
| 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 |
Osmotic(Or5h)_Roots_5_hours
|
| Organism |
Oryza sativa Japonica Group |
| Characteristics |
Tissue: 2-weeks old rice seedling Roots. 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.
|
| 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. |
|
