|
| Status |
Public on Jul 09, 2011 |
| Title |
A.thaliana 35S:GFP (Col) vs 35S:IYO-GFP (Col-0) Rep 4 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
shoot apices from 35S:GFP
|
| Organism |
Arabidopsis thaliana |
| Characteristics |
ecotype: Columbia
transgenic line: 35S:GFP
tissue: shoot apices
|
| Treatment protocol |
Shoot apices of 35S:IYO-GFP compared to shoot apices of 35S:GFP plants
|
| Growth protocol |
Arabidopsis seeds were vernalized for two days in soil and transferred to a growth chamber for 22 days under a photoperiod of 14h light/10h dark with a temperature range 20ºC/22ºC. Shoot apices were separated and immediately frozen in liquid nitrogen.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
The frozen plant tissue (~100 mg) was ground in tiny mortars dipped in liquid nitrogen. The frozen powder was carefully transfered into 1.5 ml microcentrifuge tubes using small funnels and rubber spatulas also cooled with liquid nitrogen. Total RNA was extracted using TRIzol reagent (Invitrogen, 15596-026) according to manufacturer's instructions and purified with an RNeasy mini kit (Qiagen, 74104).1 ug of total RNA was amplified via the aRNA MessageAmp II kit (Ambion, 1751)
|
| Label |
Cy3
|
| Label protocol |
5 ug of aminoallyl-labeled aRNA were resuspended in 0.1 M Na2CO3 (pH 9.0) and labeled with Hyper5 and Cy3 Mono NHS Ester (CyTMDye Post-labeling Reactive Dye Pack, Amersham). Samples were purified following the manufacturer's instructions for Megaclear TM (Ambion) and Hyper5 and Cy3 incorporation was measured using 1 ul of the probe in the Nanodrop
|
| |
|
| Channel 2 |
| Source name |
shoot apices from 35S:IYO-GFP
|
| Organism |
Arabidopsis thaliana |
| Characteristics |
ecotype: Columbia
transgenic line: 35S:IYO-GFP
tissue: shoot apices
|
| Treatment protocol |
Shoot apices of 35S:IYO-GFP compared to shoot apices of 35S:GFP plants
|
| Growth protocol |
Arabidopsis seeds were vernalized for two days in soil and transferred to a growth chamber for 22 days under a photoperiod of 14h light/10h dark with a temperature range 20ºC/22ºC. Shoot apices were separated and immediately frozen in liquid nitrogen.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
The frozen plant tissue (~100 mg) was ground in tiny mortars dipped in liquid nitrogen. The frozen powder was carefully transfered into 1.5 ml microcentrifuge tubes using small funnels and rubber spatulas also cooled with liquid nitrogen. Total RNA was extracted using TRIzol reagent (Invitrogen, 15596-026) according to manufacturer's instructions and purified with an RNeasy mini kit (Qiagen, 74104).1 ug of total RNA was amplified via the aRNA MessageAmp II kit (Ambion, 1751)
|
| Label |
Hyper5
|
| Label protocol |
5 ug of aminoallyl-labeled aRNA were resuspended in 0.1 M Na2CO3 (pH 9.0) and labeled with Hyper5 and Cy3 Mono NHS Ester (CyTMDye Post-labeling Reactive Dye Pack, Amersham). Samples were purified following the manufacturer's instructions for Megaclear TM (Ambion) and Hyper5 and Cy3 incorporation was measured using 1 ul of the probe in the Nanodrop
|
| |
|
| |
| Hybridization protocol |
The hybridization experiment was performed according to the manufacture's protocol (Agilent technologies, Agilent 60-mer oligo microarray processing protocol: Two color microarray based gene expression analysis, G4140-90050 ver 5.7)
|
| Scan protocol |
Images from Cy3 and Hyper5 channels were equilibrated and captured with a GenePix 4000B (Axon) and spots were converted into numerical data using GenPix software (Axon).
|
| Description |
Biological replicate 4 of 4. Gene expression of 35S:IYO-GFP (Col-0) vs 35S:GFP (Col-0)
|
| Data processing |
Raw intensities were background-substracted by NORMEXP method with a offset of 50. Signals (in log2 scale) were then normalized by LOWESS algorithm (intra-arrays normalization) followed by adjustment of their quantiles (inter-arrays normalization).
|
| |
|
| Submission date |
Jul 08, 2010 |
| Last update date |
Jul 09, 2011 |
| Contact name |
MAITE SANMARTIN |
| E-mail(s) |
msanmart@cnb.csic.es
|
| Organization name |
CNB-CSIC
|
| Street address |
CALLE DARWIN 3
|
| City |
MADRID |
| ZIP/Postal code |
28049 |
| Country |
Spain |
| |
|
| Platform ID |
GPL9020 |
| Series (1) |
| GSE22817 |
A mechanism for triggering cell differentiation in Arabidopsis |
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