|
| Status |
Public on Jul 31, 2013 |
| Title |
32dap-Mock treated seeds Replicate 2a |
| Sample type |
RNA |
| |
|
| Source name |
32dap_Mock2
|
| Organism |
Medicago truncatula |
| Characteristics |
tissue: seed
genetic background: R108
treatment: Mock
developmental stage: 32 days after pollination
harvest date: 12_10_29
|
| Treatment protocol |
Flowers were tagged on plants at the flower bud stage, then spray inoculated until runoff with bacterial suspensions at 1×107 CFU ml-1
|
| Growth protocol |
Medicago truncatula ssp. tricycla R108 were grown in growth chambers under 16 h of light at 22°C and 8 h of darkness at 19°C. At the flower stage, plants were transferred to a confied claimte cell at 21°C/19°C, 16h light and high (95%) relative humidity
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNAs were extracted from the frozen grinded seeds lots using the NucleoSpin Extract II, Total RNA Purification kit (Macherey-Nagel, GmbH & Co. KG, Germany) following manufacturer’s instructions
|
| Label |
Cy5
|
| Label protocol |
400ng of total RNAs were amplified using the Ambion messageAmp II (Ambion, Austin TX) following manufacturer's instructions. 5µg of each aRNAs were retrotranscribed with 400U of Superscript II reverse_transcriptase (Invitrogen Corp., Carlsbad, CA) and labelled with 1.5mmol of Cyanine_3 (Cy3) or Cyanine_5 (Cy5) (Interchim, France), then purified with NucleoSpin Gel and PCR Clean_up column kits (Macherey_Nagel, GmbH & Co. KG, Germany). Purified labeled cDNA were quantified using NanoDrop ND_1000 (Nanodrop Technologies, DE, USA). Labelled samples (30 pmol) were combined and co_hybridized to the Medtr_v1 12x135K arrays.
|
| |
|
| Hybridization protocol |
Hybridization was performed on a NimbleGen Hybridization System 4 (mix mode B) at 42° overnight. Slides were washed and dried prior to scanning.
|
| Scan protocol |
Slides were scanned at 532/635nm at 2 μm resolution and high sensitivity with a Roche_NimbleGen MS200. DEVA Software v1.2.1 was used to extract pair_data files from the scanned images.
|
| Data processing |
All statistical analyses on the gene expression data were performed using R language, version 2.5.1 (R Development Core Team, 2011) and LIMMA package (Smyth G.K., 2005) from the Bioconductor project. For the processing steps, data were normalized by the lowess method in LIMMA.
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| |
|
| Submission date |
Jul 30, 2013 |
| Last update date |
Jul 31, 2013 |
| Contact name |
Sandra Pelletier |
| E-mail(s) |
[email protected]
|
| Organization name |
INRA
|
| Lab |
IRHS
|
| Street address |
42, rue Georges Morel - BP 60057
|
| City |
BEAUCOUZE |
| ZIP/Postal code |
49045 |
| Country |
France |
| |
|
| Platform ID |
GPL16373 |
| Series (1) |
| GSE49347 |
Transmission of xanthomonads to Medicago truncatula seeds |
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