|
| Status |
Public on Dec 31, 2013 |
| Title |
oldDC -Fe_rep2 |
| Sample type |
RNA |
| |
|
| Source name |
vascular bundle from old leaf at the lowest node grown under Fe-deficient condition
|
| Organism |
Oryza sativa |
| Characteristics |
tissue: vascular bundle from old leaf at the lowest node
growth condition: Fe-deficient
|
| Treatment protocol |
Fe deficiency was initiated at day 27 after germination by omitting Fe(III)–EDTA from the culture medium, and plants were harvested after 7 days. For Cd stress, 10 μM CdCl2 was added to the hydroponic culture on day 33 after germination and plants were harvested after 24 h.
|
| Growth protocol |
Rice was grown hydroponically under 14-h/10-h light/dark cycles and at 30/25°C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
The 0.5 cm to 1.5 cm of the root tip and DC were fixed in 75% ethanol/25% acetate and 100% acetone, respectively. Paraffin-embedding was followed by the microwave method. Serial paraffin sections were prepared at 10–20 μm in thickness, and they were put onto a glass slide. Laser Microdissection was performed using the Veritas Laser Microdissection System LCC1704 (Molecular Devices). Total RNA was extracted from cells prepared by Laser Microdissection with a Pico-PureTM RNA isolation kit (Molecular Devices) according to the manufacturer’s protocol. The extracted total RNA was quantified with a Quant-iTTM RiboGreen RNA reagent and kit (Invitrogen, Carlsbad, CA) according to the manufacturer’s protocol. The quality of total RNA was assessed using an RNA 6000 Pico kit on the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA). RNA integrity was judged by RNA Integrity Number (RIN), which was calculated with 2100 Expert Software (Agilent, version B.02.02, eukaryote total RNA pico mode).
|
| Label |
Cy3
|
| Label protocol |
One-color spikemix was added to the total RNA prior to the labeling reaction, and labeling was performed using a Quick Amp Labeling Kit, One-Color (Agilent Technologies) in the presence of cyanine-3 (Cy3)-CTP, according to the modified manufacturer’s protocol. The Cy3-labeled cRNA was purified by RNeasy Mini Kit (Qiagen), and the quantity was examined with a NanoDrop ND-1000 UV-VIS spectrophotomer (NanoDrop Technologies).
|
| |
|
| Hybridization protocol |
Cy3-labeled cRNA was fragmented and hybridized on a slide of rice 4 • 44K microarray RAP-DB (G2519F#15241; Agilent Technologies) at 65°C for 17 h. Hybridization and washing of the hybridized slide were performed according to the manufacturer’s instructions.
|
| Scan protocol |
Slides were scanned on an Agilent G2505B DNA microarray scanne.
|
| Description |
Gene expression of vascular bundle from old leaf at the lowest node grown under Fe-deficient condition
|
| Data processing |
The scanned images were analyzed with FEATURE EXTRACTION 10.5.1.1 (Agilent Technologies) to obtain background correction of the Cy3 raw signals.
|
| |
|
| Submission date |
Aug 31, 2012 |
| Last update date |
Dec 31, 2013 |
| Contact name |
Yuko Ogo |
| E-mail(s) |
[email protected]
|
| Phone |
+81 29 838 8681
|
| Organization name |
NARO
|
| Street address |
3-1-3 Kannondai
|
| City |
Tsukubashi |
| State/province |
Ibaraki |
| ZIP/Postal code |
305-0035 |
| Country |
Japan |
| |
|
| Platform ID |
GPL6864 |
| Series (1) |
| GSE40549 |
Spatial transcriptomes of iron-deficient and cadmium-stressed rice |
|
