|
| Status |
Public on Jul 01, 2013 |
| Title |
AG amiRNA knockdown 3 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
LhG4 GR set3
|
| Organism |
Arabidopsis thaliana |
| Characteristics |
time: 0 d
treatment: No Dexamethasone
tissue: Whole infloresence
sample type: Whole infloresence, no Dexamethasone treatment
genotype: LhG4-GR_N6XpOp-AmiRNA-AG (ler)
genetic background: ler
|
| Treatment protocol |
For all experiments, ~4 week-old plants were used. For experiments using the FIS (35S:AP1-GR ap1-1 cal-1 plants), flower development was induced using a solution containing 10 μM dexamethasone (Sigma-Aldrich), 0.01% (v/v) ethanol and 0.015% (v/v) Silwet L-77 (De Sangosse) – as described in Wellmer F, et al. (2006) Genome-Wide Analysis of Gene Expression during Early Arabidopsis Flower Development. PLoS Genetics 2(7):e117.
|
| Growth protocol |
Plants were grown on a soil:vermiculite:perlite (3:1:1) mixture at 20 degrees under constant illumination with cool white fluorescent light
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated from tissue samples using the Plant Total RNA kit (Sigma-Aldrich), according to the manufacturer's instructions
|
| Label |
Cy5
|
| Label protocol |
500ng of total RNA were primed with 0.6µl of T7 DNA primer at 65°C for 10 min, then reverse transcribed at 40°C for 2 hrs in the presence of 300 U MMLV Reverse Transcriptase and 10mM dNTP mix, with Cy3-label and Cy5- label.
|
| |
|
| Channel 2 |
| Source name |
LhG4 GR set3
|
| Organism |
Arabidopsis thaliana |
| Characteristics |
sample type: Whole infloresence, Dexamethasone treatment
time: 0 d
treatment: Dexamethasone
tissue: Whole infloresence
genotype: LhG4-GR_N6XpOp-AmiRNA-AG (ler)
genetic background: ler
|
| Treatment protocol |
For all experiments, ~4 week-old plants were used. For experiments using the FIS (35S:AP1-GR ap1-1 cal-1 plants), flower development was induced using a solution containing 10 μM dexamethasone (Sigma-Aldrich), 0.01% (v/v) ethanol and 0.015% (v/v) Silwet L-77 (De Sangosse) – as described in Wellmer F, et al. (2006) Genome-Wide Analysis of Gene Expression during Early Arabidopsis Flower Development. PLoS Genetics 2(7):e117.
|
| Growth protocol |
Plants were grown on a soil:vermiculite:perlite (3:1:1) mixture at 20 degrees under constant illumination with cool white fluorescent light
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated from tissue samples using the Plant Total RNA kit (Sigma-Aldrich), according to the manufacturer's instructions
|
| Label |
Cy3
|
| Label protocol |
500ng of total RNA were primed with 0.6µl of T7 DNA primer at 65°C for 10 min, then reverse transcribed at 40°C for 2 hrs in the presence of 300 U MMLV Reverse Transcriptase and 10mM dNTP mix, with Cy3-label and Cy5- label.
|
| |
|
| |
| Hybridization protocol |
Fragmentation buffer and Hybridization Buffer HI-RPM (Agilent Gene Expression Hybridization Kit) were added and samples were applied to microarrays enclosed in Agilent SureHyb-enabled hybridization chambers and hybridized for 16 hrs at 65°C. After hybridization, slides were washed sequentially according to the manufacturer's instruction.
|
| Scan protocol |
Scanned on an Agilent G2565AA scanner. Images were quantified using Agilent Feature Extraction Software (v 9.5.3.1).
|
| Description |
Biological Replicate 3: several plants (approx 20-25) pooled per replicate and genotype
|
| Data processing |
Agilent Feature Extraction Software (v 9.5.3.1) was used for background subtraction and LOWESS normalization
|
| |
|
| Submission date |
Mar 06, 2013 |
| Last update date |
Jul 01, 2013 |
| Contact name |
Pat Ryan |
| E-mail(s) |
pat.ryan12@gmail.com
|
| Organization name |
TCD
|
| Street address |
College Green
|
| City |
Dublin |
| ZIP/Postal code |
na |
| Country |
Ireland |
| |
|
| Platform ID |
GPL9984 |
| Series (2) |
| GSE44919 |
Control of Reproductive Floral Organ Identity Specification in Arabidopsis by the C Function Regulator AGAMOUS (expression) |
| GSE45939 |
Control of Reproductive Floral Organ Identity Specification in Arabidopsis by the C Function Regulator AGAMOUS |
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