|
| Status |
Public on Aug 05, 2018 |
| Title |
Wild type mechanical damage vs PDKO3 mechanical damage 1 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
WT rosette leaves mechanical damaged
|
| Organism |
Arabidopsis thaliana |
| Characteristics |
treatment: mechanical damaged
age: day 22 (growth stage 3)
tissue: rosette leaves
ecotype: Columbia 0
genotype: WT
|
| Treatment protocol |
Mechanical damage: made by pattern wheel mimicking herbivore; Herbivore wounding: Spodoptera littoralis feeding
|
| Growth protocol |
Arabidopsis plants (Col-0 and PDKO3) were grown as a single plant in plastic pots filled with soil. Growth chamber was set up at 23°C 16h in the light (160 μE m-2 s-1) and 8 in the dark for 22 days
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extracted from rosette leaves using Agilent Plant RNA Isolation Mini kit including RNAse-free DNase (Qiagen) treatment
|
| Label |
Cy3
|
| Label protocol |
500 ng of total RNA were separately reverse-transcribed into double-strand cDNAs by the Moloney murine leukemia virus reverse transcriptase (MMLV-RT) and amplified for 2 h at 40°C using the Agilent Quick Amp Labelling Kit, two-color (Agilent Technologies). Subsequently, cDNAs were transcribed into antisense cRNA and labeled with either Cy3-CTP or Cy5-CTP fluorescent dyes for 2 h at 40°C following the manufacturer’s protocol. Cyanine-labeled cRNAs were purified using RNeasy Minikit (Qiagen). Purity and dye incorporation were assessed with the NanoDrop ND-1000 UV-VIS Spectrophotometer (Thermo Fisher Scientific) and the Agilent 2100 Bioanalyzer (Agilent Technologies).
|
| |
|
| Channel 2 |
| Source name |
PDKO3 rosette leaves mechanical damaged
|
| Organism |
Arabidopsis thaliana |
| Characteristics |
treatment: mechanical damaged
ecotype: Columbia 0
age: day 22 (growth stage 3)
tissue: rosette leaves
genotype: mutated in 3 plasmodesmata genes
|
| Treatment protocol |
Mechanical damage: made by pattern wheel mimicking herbivore; Herbivore wounding: Spodoptera littoralis feeding
|
| Growth protocol |
Arabidopsis plants (Col-0 and PDKO3) were grown as a single plant in plastic pots filled with soil. Growth chamber was set up at 23°C 16h in the light (160 μE m-2 s-1) and 8 in the dark for 22 days
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extracted from rosette leaves using Agilent Plant RNA Isolation Mini kit including RNAse-free DNase (Qiagen) treatment
|
| Label |
Cy5
|
| Label protocol |
500 ng of total RNA were separately reverse-transcribed into double-strand cDNAs by the Moloney murine leukemia virus reverse transcriptase (MMLV-RT) and amplified for 2 h at 40°C using the Agilent Quick Amp Labelling Kit, two-color (Agilent Technologies). Subsequently, cDNAs were transcribed into antisense cRNA and labeled with either Cy3-CTP or Cy5-CTP fluorescent dyes for 2 h at 40°C following the manufacturer’s protocol. Cyanine-labeled cRNAs were purified using RNeasy Minikit (Qiagen). Purity and dye incorporation were assessed with the NanoDrop ND-1000 UV-VIS Spectrophotometer (Thermo Fisher Scientific) and the Agilent 2100 Bioanalyzer (Agilent Technologies).
|
| |
|
| |
| Hybridization protocol |
Hybridization buffer (Agilent Gene Expression Hybridization Kit ) were added, and samples were applied to microarrays enclosed in Agilent SureHyb-enabled hybridization chambers. After hybridization, slides were washed sequential
|
| Scan protocol |
Scanned on an Agilent G2505B scanner.
Images were quantified using Agilent Feature Extraction Software (v 9.5.1.1)
|
| Description |
WT mechanical damaged leaves (control) vs PDKO3 mechanical damaged leaves (treatment). Harvested 2 h after the treatments
|
| Data processing |
R package limma (v 3.4.4) was used for background subtraction and LOESS normalization.
|
| |
|
| Submission date |
Aug 08, 2011 |
| Last update date |
Aug 05, 2018 |
| Contact name |
Cinzia Margherita Bertea |
| E-mail(s) |
[email protected]
|
| Phone |
+39-011-6706361
|
| Fax |
+39-011-6706364
|
| Organization name |
University of Turin
|
| Department |
Department of Life Sciences and Systems Biology
|
| Lab |
Plant Physiology Unit
|
| Street address |
via Quarello 11/A
|
| City |
Turin |
| State/province |
To |
| ZIP/Postal code |
10135 |
| Country |
Italy |
| |
|
| Platform ID |
GPL12621 |
| Series (1) |
| GSE31268 |
Arabidopsis thaliana wild type and PDKO3 mutant mechanical damage (MD) vs herbivore wounding (HW) |
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