|
| Status |
Public on Jan 01, 2012 |
| Title |
eid3_45 min after red-light pulse_3 |
| Sample type |
RNA |
| |
|
| Source name |
eid3 mutant seedlings 45 min after red light pulse 3
|
| Organism |
Arabidopsis thaliana |
| Characteristics |
genotype: eid3 mutant
light treatment: 2 min red-light pulse + 45 min darkness
|
| Treatment protocol |
4-days-old dark-grown seedlings were either kept in darkness or irradiated for 2 min with red-light (30 µmol/m2s) and harvested after further 45 min in darkness.
|
| Growth protocol |
Arabidopsis thaliana seeds were sterilized with ethanol and sown on 1/2 MS Agar plates covered with filter paper. After stratification for 3 days at 6°C imbibed seeds were treated with red light (30 µmol/m2s) for 2 h at 21°C to induce germination. Seedlings were grown for 4 days at 21°C in darkness.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Harvested seedlings were immediately frozen in liquid nitrogen. The material was homogenized in eppendorf-tubes together with 7 glass beads (2 mm size) in frozen condition in a silamat homogenizer (vivadent ivoclar) for 2x 10 s. Total RNA was extracted using the Quiagen Plant RNeasy Mini Kit followong the manufacturer´s recommendations. The protocol includes an on-column DNase (Quiagen) digestion step. RNA was quantified using a NanoDrop-1000 spectrophotometer (peqlab Biotechnology GmbH) and quality was estimated on an Agilent 2100 Bioanalyzer. The RNA integrity number index was calculated for each sample using Agilent 2100 Expert software. Only RNAs with integrity number >8.0 were further processed.
|
| Label |
Cy3
|
| Label protocol |
0.5 µg of total RNA was reverse transcribed into cDNA and labeled with Cy3 using the one-color Quick Amp Labeling Kit (Agilent) following Agilent´s instructions.
|
| |
|
| Hybridization protocol |
Hybridization was performed with 1.5 µg of labeled cDNA per array at a temperature of 65°C overnight, according to the Agilent protocol.
|
| Scan protocol |
scanned on Agilent Technologies Scanner G2505C at a resolution of 5 μm.
|
| Description |
Gene expression 45 min after a red-light pulse in eid3 mutant seedlings.
|
| Data processing |
Agilent Feature Extraction Software 10.5.1.1 was used to process and analyze array images. The software returns a series of spot quality measures to evaluate the quality and reliability of spot intensity estimates. The raw data were analyzed using GeneSpring GX 10.0 (normalization: shift to 75.0 percentile, baseline transformation: median of all samples). The normalized data were filtered to exclude probes flagged “absent” in all samples.
|
| |
|
| Submission date |
Nov 21, 2011 |
| Last update date |
Jan 01, 2012 |
| Contact name |
Cornelia Klose |
| E-mail(s) |
cornelia.klose@biologie.uni-freiburg.de
|
| Organization name |
Albert-Ludwigs-University Freiburg
|
| Department |
Institute for Biology 2/ Botany
|
| Lab |
Thomas Kretsch
|
| Street address |
Schänzlestr. 1
|
| City |
Freiburg |
| ZIP/Postal code |
79104 |
| Country |
Germany |
| |
|
| Platform ID |
GPL9020 |
| Series (1) |
| GSE33856 |
Arabidopsis thaliana seedlings:early light regulated gene expression in Ler wildtype versus eid3 mutant |
|
