|
| Status |
Public on Jan 15, 2015 |
| Title |
Apex-2N-rep3 |
| Sample type |
RNA |
| |
|
| Source name |
B. napus Apex - 2 mM N - rep3
|
| Organism |
Brassica napus |
| Characteristics |
cultivar: Apex
nitrogen efficiency status: N efficient
tissue: leaf number 3
treatment: N standard control
|
| Treatment protocol |
Leaf senescence was induced by (i) N starvation (0.1 mM N as Ca(NO3)2 and 1 mM CaSO4), (ii) shading of leaf 3 (counted from bottom to top of the plant), and (iii) detaching leaf 3 and incubating it in deionized water.
|
| Growth protocol |
Seven days old seedlings were hydroponically pre-cultured for 28 days at optimal N supply in a continuously aerated nutrient solution composed of 2 mM N [9:1 ratio of Ca(NO3)2 and (NH4)2SO4], 500 µM K2S04, 250 µM KH2PO4, 325 µM MgSO4, 50 µM NaCl, 8 µM H3BO3, 0.4 µM MnSO4, 0.4 µM ZnSO4, 0.4 µM CuSO4, 0.1 µM MoNa2O4, 40 µM Fe-EDDHA and 10 µM C2H4N4.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from 500 mg frozen and ground tissue according to Verwoerd et al. (1989. Nucleic Acids Res. 17, 2362) and purified with the RNeasy Mini Kit (Qiagen, Hilden, Germany). RNA was photometrically quantified with a NanoPhotometer (Implen, München, Germany) and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA).
|
| Label |
Cy3
|
| Label protocol |
Cy3-labeled cRNA was synthesized with the Low Input Quick Amp Labeling Kit, one-color (Agilent Technologies catalog number 5190-2305) according to the manufacturer's instructions.
|
| |
|
| Hybridization protocol |
0.6 ug of Cy3-labelled cRNA was fragmented in 1x Agilent fragmentation buffer (Agilent Technologies catalog number 5188-5242) and 2x Agilent blocking agent (Agilent Technologies catalog number 5188-5242) following the manufacturers instructions. Hybridization was performed in a Agilent hybridization oven (Agilent Technologies catalog number G2545-90002) at standard conditions as recommended by the manufacturer. Subsequently, microarrays were washed 1 minute with GE Wash Buffer 1 at ambient temperature and 1 minute with GE Wash buffer 2 (Agilent Technologies catalog number 5188-5327) at 37°C.
|
| Scan protocol |
Microarrays were scanned immediately after washing with an Agilent High-Resolution Scanner (G2505C) using default settings.
|
| Description |
Gene expression in control leaves with standard N supply
|
| Data processing |
The images were processed with the Agilent Feature Extraction software 10.1 using default settings (protocol GE1_105_Dec08 (Read Only) and GridName US91803681_252963410002_S01_grid (from grid_file)). The resulting processed signals of all individual hybridizations were quantile-normalized using the ranked median quantiles according to Bolstad et al. (2003. Bioinformatics 19, 185-193).
|
| |
|
| Submission date |
Aug 05, 2014 |
| Last update date |
Jan 15, 2015 |
| Contact name |
Reinhard Kunze |
| E-mail(s) |
reinhard.kunze@fu-berlin.de
|
| Phone |
+49 30 838 55802
|
| Organization name |
Freie Universität Berlin
|
| Department |
Institute of Biology
|
| Lab |
Plant Molecular Genetics
|
| Street address |
Albrecht-Thaer-Weg 6
|
| City |
Berlin |
| State/province |
Berlin |
| ZIP/Postal code |
14195 |
| Country |
Germany |
| |
|
| Platform ID |
GPL19044 |
| Series (1) |
| GSE60108 |
Nitrogen starvation and cultivar-specific leaf senescence in winter oilseed rape (Brassica napus) |
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