|
| Status |
Public on Sep 01, 2010 |
| Title |
Day 0, Normoxia, Susceptible oysters, pool 3 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Pooled Digestive Gland, Susceptible Oysters, Starting point
|
| Organism |
Magallana gigas |
| Characteristics |
oyster line: Susceptible Oysters
tissue: Digestive Gland
|
| Treatment protocol |
Hypoxia exposure maintained for 20 d. The oxygen levels were reduced by bubbling nitrogen to obtain 2 mg O2 L-1 (30% oxygen saturation at 20°C)
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extracted using Extract-all Reagent (Eurobio) manufacturer's instructions
|
| Label |
Cy5
|
| Label protocol |
Five μg of total RNA were directly labeled by reverse transcription using the Direct ChipShot Direct Labeling and Clean-Up Kit (Promega) according to the manufacturer’s recommendations.
|
| |
|
| Channel 2 |
| Source name |
Total RNA from pooled entire wild oysters
|
| Organism |
Magallana gigas |
| Characteristics |
oyster line: Wild Oysters
tissue: All Tissues
|
| Treatment protocol |
Hypoxia exposure maintained for 20 d. The oxygen levels were reduced by bubbling nitrogen to obtain 2 mg O2 L-1 (30% oxygen saturation at 20°C)
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extracted using Extract-all Reagent (Eurobio) manufacturer's instructions
|
| Label |
Cy3
|
| Label protocol |
Five μg of total RNA were directly labeled by reverse transcription using the Direct ChipShot Direct Labeling and Clean-Up Kit (Promega) according to the manufacturer’s recommendations.
|
| |
|
| |
| Hybridization protocol |
The slides were filled with a pre-hybridization buffer (Chip Spread buffer containing 4× SSC and 0.2% SDS) for 1 h at 42°C. Hybridization was conducted overnight at 42°C in the automatic station according to the manufacturer’s instructions. After hybridization, the arrays were washed twice with Ribowash solution (0.1 M Tris, 0.05 M EDTA, and 0.4 M NaCl) and once with 0.1× SSC, and finally centrifuged (6,000 rpm, 15 s, room temperature) for drying
|
| Scan protocol |
Microarray slides were scanned using a Genepix 4000B microarray scanner (Axon Instruments Inc.) with standard dual laser excitation at 532 nm (17 mW) and 635 nm (10 mW) according to the following parameters: Cy5 Photo Multiplier Tube (PMT) 550 and Cy3 PMT 590. Images (16-bit TIF) were analyzed using GenePix pro 5.1 software (Axon Instruments Inc.) according to the manufacturer’s instructions
|
| Data processing |
Microarray data were initially processed using the language R/BioConductor (R Development Core Team 2008). LOESS normalization and background correction were performed with the limma package. Missing (NA) values where calculated with an application of the TMev Software (A.I. Saeed, N.K. Bhagabati, J.C. Braisted, W. Liang, V. Sharov, E.A. Howe, et al., TM4 Microarray Software Suite, in: DNA Microarrays, Part B: Databases and Statistics, Academic Press, 2006: pp. 134-193).
|
| |
|
| Submission date |
Aug 31, 2010 |
| Last update date |
Aug 31, 2010 |
| Contact name |
Rossana Sussarellu |
| E-mail(s) |
rossana.sussarellu@ifremer.fr
|
| Organization name |
Ifremer
|
| Department |
Ressources Biologiques et Environnement
|
| Lab |
Laboratoire d'Ecotoxicologie
|
| Street address |
Rue de l'Ile d'Yeu
|
| City |
Nantes |
| ZIP/Postal code |
44300 |
| Country |
France |
| |
|
| Platform ID |
GPL8639 |
| Series (1) |
| GSE23883 |
Transcriptomic response of the Pacific oyster Crassostrea gigas to hypoxia |
|
