|
| Status |
Public on May 25, 2012 |
| Title |
IC-547557 Biological replicate 2 |
| Sample type |
RNA |
| |
|
| Source name |
IC-547557 Low-N tolerant
|
| Organism |
Oryza sativa |
| Characteristics |
tissue: whole plant
genotype: IC-547557 Low-N tolerant
|
| Treatment protocol |
0.01mM NH4NO3 was used as low N condition to grow genotypes IC-547557 and Vivek Dhan.
|
| Growth protocol |
Seeds were germinated in the dark at 27°C on blotting paper saturated with deionized water. After 96 h, seedlings were transferred to a hydroponic system placed in a growth chamber with a day/night regime of 16/8 h and a PPFD of 200 µmol m-2 s-1 at plant level, with a temperature of 22°C in the dark and 26°C in the light and with a relative humidity of 65%. Seedlings were grown using the nutrient medium containing 2 mM K2SO4, 2 mM MgSO4, 1 mM CaCl2, 0.3 mM NaH2PO4, 40 µM Fe-EDTA, 9 µM MnCl2, 25 µM Na2MoO4, 20 µM H3BO3, 1.5 µM ZnSO4, and 1.5 µM CuSO4. The nitrogen in the form of NH4NO3 was maintained as 0.01 mM (low-N condition). The pH of the nutrient solutions was adjusted to 6.0 and the solutions were changed every three days. 21 day old seedlings of eachIC-547557 and Vivek Dhan was selected for miRNA array.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Whole plant samples were flesh freeze in liquid nitrogen. For miRNA array procedure total RNA was isolated using TRIZOL® reagent (Invitrogen, USA) with additional isopropanol overnight precipitation at -20 °C. Concentration of isolated RNA was determined using iTTM RiboGreen RNA assay Kit (Invitrogen, USA).
|
| Label |
Biotin
|
| Label protocol |
Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 500 ng total RNA (FlashTag® Biotin HSR RNA Labeling Kit for Affymetrix® GeneChip® miRNA Arrays from GENISPHERE)
|
| |
|
| Hybridization protocol |
For array preparation (GeneChip miRNA_2.0 Array) 21.5 µl of biotin labeled samples were incubated with array hybridization cocktail which includes 2× hybridization buffer, 27.5% formamide, dimethyl sulphoxide (DMSO), 20× eukaryotic hybridization controls and control Oligonucleotide (from GeneChip eukaryotic hybridization control kit). 100 µl of this cocktail was injected into arrays and incubated in hybridization oven at 48 °C and 60 rpm for 16 h. This cocktail was washed and stained with Fluidics Station 450. Filled array was then scanned using Gene chip scanner 3000 7G.
|
| Scan protocol |
Filled array was then scanned using Gene chip scanner 3000 7G
|
| Description |
microRNA expression data from whole rice plant 21 day old seedling grown under low-N condition.
|
| Data processing |
Probe specific signal detection call is based on Wilcoxon Rank sum test of the miRNA probes compared to the distribution of signal from GC content. Filtering was done to remove unimportant (non-rice) as well as unreliable data. After scanning data summarization, normalization and quality control was done through miRNA QC Tool software version 1.1.1.0, available at www.affymetrix.com.
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| |
|
| Submission date |
May 24, 2012 |
| Last update date |
May 25, 2012 |
| Contact name |
ALTAF AHMAD |
| E-mail(s) |
aahmed@jamiahamdard.ac.in
|
| Phone |
911126059688
|
| Fax |
91112609663
|
| Organization name |
Jamia Hamdard (Hamdard University)
|
| Department |
Department of Botany
|
| Lab |
Molecular Ecology Laboratory
|
| Street address |
Faculty of Science
|
| City |
New Delhi |
| State/province |
Delhi |
| ZIP/Postal code |
110062 |
| Country |
India |
| |
|
| Platform ID |
GPL14613 |
| Series (1) |
| GSE38213 |
Identification and comparative analysis of microRNAs associated with low-N tolerance in rice genotypes |
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