|
| Status |
Public on Oct 26, 2011 |
| Title |
Root cap_rep2 |
| Sample type |
RNA |
| |
|
| Source name |
Root cap
|
| Organism |
Oryza sativa Japonica Group |
| Characteristics |
cultivar: Nipponbare
tissue: root
developmental stage: root cap
|
| Treatment protocol |
The crown roots of 10-day-old seedlings with length more than 5 cm were used for the gene expression profiling. The samples were fixed in 99.5% cold ethanol or 100% cold acetone. Sample fixation and paraffin-embedding were performed using the Microwave Processor (Energy Beam Sciences, East Granby, CT). The paraffin-embedded samples were cut into 12-20 µm thick sections using a microtome (Leica RM2255) and used for laser microdissection with the Veritas Laser Microdissection System LCC1704 (Arcturus).
|
| Growth protocol |
Rice seeds were sterilized with 70% (v/v) ethanol solution and 1% (v/v) sodium hypochlorite solution, imbibed in distilled water in the dark at 25°C for 2 days, and transferred onto a nylon net floated in distilled water in a growth chamber (60% humidity; 14 h light period at 30°C and 10 h dark period at 25°C). After 3 days of germination, the seedlings were transferred to a nutrient solution (based on Yoshida nutrient solution; pH 5.5) and allowed to grow for another 7 days. The pH of the nutrient solution was adjusted using 1N NaOH and maintained using MES (buffer), and the solution was renewed every 2 days.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from each sample with a Pico-Pure RNA isolation kit (Arcturus) according to the manufacturer’s protocol. The quantity and quality of the obtained RNAs were checked with an Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA, USA).
|
| Label |
Cy3
|
| Label protocol |
One-color spike-mix 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 cyanine-3-labeled cRNA was purified by the RNeasy Mini Kit (Qiagen), and the cRNA yield was determined using the NanoDrop ND-1000 UV-VIS spectrophotomer (NanoDrop).
|
| |
|
| Hybridization protocol |
At least 1,100 ng Cy3-labeled cRNA was fragmented and hybridized on a slide of rice 4x44K microarray RAP-DB (Agilent; G2519F#15241) at 65℃ for 17 hours. 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 scanner.
|
| Description |
RXP4001_002
Gene expression of root cap.
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 10.5.1.1 (Agilent) using default parameters to obtain background subtracted and spatially detrended processed signal intensities. The processed signal intensities were used for 75th percentile normalization and log2 transformation with GeneSpring GX11 (Agilent).
|
| |
|
| Submission date |
Jun 21, 2011 |
| Last update date |
Oct 27, 2011 |
| Contact name |
Yutaka Sato |
| E-mail(s) |
satoyu@affrc.go.jp
|
| Organization name |
National Institute of Agrobiological Sciences
|
| Street address |
2-1-2 Kannondai
|
| City |
Tsukuba |
| State/province |
Ibaraki |
| ZIP/Postal code |
305-8602 |
| Country |
Japan |
| |
|
| Platform ID |
GPL6864 |
| Series (1) |
| GSE30136 |
Comprehensive gene expression profiling of the rice root system based on developmental stages and tissue types |
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