|
| Status |
Public on Apr 24, 2013 |
| Title |
Uninjured VHb leaves positioned below herbivory exposure 1 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Control treatment, VHb B leaf
|
| Organism |
Populus tremula x Populus tremuloides |
| Characteristics |
line: V617/45, generated via Agrobacterium mediated gene transfer using pBVHb
tissue: leaf
leaf type: below leaf, LPI 11
plant number: 38
treatment: none
|
| Treatment protocol |
The saplings of wt and VHb lines grown 1.5 months under standard greenhouse conditions were used in the experiments. In the herbivory treatment, one larvae of Conistra vaccinii L. (Lepidoptera: Noctuidae) was placed on the leaf inside the veil cloth bag for 24 hours, which after samples were collected to liquid nitrogen.
|
| Growth protocol |
The lines were multiplicated on liquid MS medium (full strength of C10H12FeN2NaO8; half strength of other micro and macro nutrients; 2.22 μM BA and 2.85 μM IAA) in RITA® temporary immersion containers (Vitropic, Saint-Mathieu-de-Tréviers, France) under 16:8 h light/dark photoperiod (110-130 μmol m-2s-1) at 22 ºC. Saplings were rooted on MS without plant growth regulators, acclimated in the Botanical Gardens of the University of Oulu (65°03'N, 25°27'E), and potted to limed and fertilised soil, and grown under standard greenhouse conditions.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
The total RNA was extracted as described by Jaakola et al. (Mol. Biotechnol. 19 (2001) 201-203) except for the first centrifugation step which was omitted.
|
| Label |
Cy5
|
| Label protocol |
The SuperScript III reverse transcriptase (Invitrogen, Carlsbad, CA, USA) was used to synthesize cDNAs in accordance with the manufacturer's instructions and the instructions of the 3DNA Array 50 Expression Array Detection Kit For Microarrays (Genisphere Inc, Hatfield, PA, USA).
|
| |
|
| Channel 2 |
| Source name |
Herbivory treatment, VHb B leaf
|
| Organism |
Populus tremula x Populus tremuloides |
| Characteristics |
line: V617/45, generated via Agrobacterium mediated gene transfer using pBVHb
tissue: leaf
leaf type: below leaf, LPI 11
plant number: 25
treatment: herbivore (chestnut moth larvae)
|
| Treatment protocol |
The saplings of wt and VHb lines grown 1.5 months under standard greenhouse conditions were used in the experiments. In the herbivory treatment, one larvae of Conistra vaccinii L. (Lepidoptera: Noctuidae) was placed on the leaf inside the veil cloth bag for 24 hours, which after samples were collected to liquid nitrogen.
|
| Growth protocol |
The lines were multiplicated on liquid MS medium (full strength of C10H12FeN2NaO8; half strength of other micro and macro nutrients; 2.22 μM BA and 2.85 μM IAA) in RITA® temporary immersion containers (Vitropic, Saint-Mathieu-de-Tréviers, France) under 16:8 h light/dark photoperiod (110-130 μmol m-2s-1) at 22 ºC. Saplings were rooted on MS without plant growth regulators, acclimated in the Botanical Gardens of the University of Oulu (65°03'N, 25°27'E), and potted to limed and fertilised soil, and grown under standard greenhouse conditions.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
The total RNA was extracted as described by Jaakola et al. (Mol. Biotechnol. 19 (2001) 201-203) except for the first centrifugation step which was omitted.
|
| Label |
Cy3
|
| Label protocol |
The SuperScript III reverse transcriptase (Invitrogen, Carlsbad, CA, USA) was used to synthesize cDNAs in accordance with the manufacturer's instructions and the instructions of the 3DNA Array 50 Expression Array Detection Kit For Microarrays (Genisphere Inc, Hatfield, PA, USA).
|
| |
|
| |
| Hybridization protocol |
Hybridizations and post-washings were conducted following the instructions of 3DNA Array 50 Expression Array Detection Kit (Genisphere Inc).
|
| Scan protocol |
Slides were scanned at 633 nm and 543 nm at 98 % laser power, at a resolution of 10 µm with varying PMT values with ScanArray® Gx PLUS (Microarray Scanner, PerkinElmer, Waltham, MA, USA). The fixed-circle method was used in the quantification of spots, and the total-algorithm in the normalization was conducted with the ScanArray Express software (PerkinElmer).
|
| Description |
VHb_B_rep1
Replicate 1.
|
| Data processing |
The statistical processing of the data was conducted separately on the two lines and different leaf samples by using identical procedures. The low quality spots were downweighted; these included spots with the sum of median pixel intensities less than 300 and spots having the feature pixel intensity less than 55 % above one standard deviation of the background pixel intensity in both wavelengths. The data was normalized by using print-tip loess within and Aquantile (Aq) between array normalizations.
|
| |
|
| Submission date |
Apr 23, 2013 |
| Last update date |
Apr 24, 2013 |
| Contact name |
Suvi Tuulia Sutela |
| E-mail(s) |
suvi.sutela@oulu.fi
|
| Organization name |
University of Oulu
|
| Department |
Biology
|
| Street address |
P.O. Box 3000
|
| City |
Oulu |
| ZIP/Postal code |
90014 |
| Country |
Finland |
| |
|
| Platform ID |
GPL17056 |
| Series (1) |
| GSE46331 |
24-hour herbivory exposure of VHb-expressing Populus tremula x Populus tremuloides |
|
