|
| Status |
Public on Aug 17, 2011 |
| Title |
Cumulus enclosed vs. cumulus denuded oocytes slide13903776_CEO2cy5-CDO2cy3 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
cow oocyte, mature metaphase-II, CDO replicate2
|
| Organism |
Bos taurus |
| Characteristics |
maturation method: cumulus enclosed oocyte maturation
cell type: in vitro matured cumulus enclosed oocyte
|
| Treatment protocol |
CEO were mechanically detached from cumulus and oocytes from both CEO and CDO groups were collected and then kept frozen at -80°C in RNAlater until RNA extraction.
|
| Growth protocol |
Bovine ovaries were collected from a slaughterhouse; 3-6 mm antral follicles were aspirated. CEO and CDO oocytes were then matured during 22h in enriched TCM199 medium (Sigma).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Groups of 25 CDO and CEO oocytes were extracted using Picopure RNA isolation kit (Alphelys, Plaisir, France) according to the manufacturer’s instructions. Total RNA were subsequently amplified using RiboAmp Plus kit (Alphelys, Plaisir, France) following the manufacturer’s instructions and aRNA produced were validated using the Bioanalyzer 2100 RNA 6000 nanochip (Agilent Technologies).
|
| Label |
Cy5
|
| Label protocol |
2 µg of each sample were labelled using the ULS aRNA labelling kit (Kreatech, Amsterdam, Netherlands) according manufacturer’s instructions. A dye swap approach was used by labelling CEO and CDO alternatively by cyanine 3 and 5.
|
| |
|
| Channel 2 |
| Source name |
cow oocyte, mature metaphase-II, CEO replicate2
|
| Organism |
Bos taurus |
| Characteristics |
maturation method: cumulus denuded oocyte maturation
cell type: in vitro matured cumulus denuded oocyte
|
| Treatment protocol |
CEO were mechanically detached from cumulus and oocytes from both CEO and CDO groups were collected and then kept frozen at -80°C in RNAlater until RNA extraction.
|
| Growth protocol |
Bovine ovaries were collected from a slaughterhouse; 3-6 mm antral follicles were aspirated. CEO and CDO oocytes were then matured during 22h in enriched TCM199 medium (Sigma).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Groups of 25 CDO and CEO oocytes were extracted using Picopure RNA isolation kit (Alphelys, Plaisir, France) according to the manufacturer’s instructions. Total RNA were subsequently amplified using RiboAmp Plus kit (Alphelys, Plaisir, France) following the manufacturer’s instructions and aRNA produced were validated using the Bioanalyzer 2100 RNA 6000 nanochip (Agilent Technologies).
|
| Label |
Cy3
|
| Label protocol |
2 µg of each sample were labelled using the ULS aRNA labelling kit (Kreatech, Amsterdam, Netherlands) according manufacturer’s instructions. A dye swap approach was used by labelling CEO and CDO alternatively by cyanine 3 and 5.
|
| |
|
| |
| Hybridization protocol |
50 pmol of each condition (CEO and CDO) were then fragmented for 15 minutes at 70°C using the RNA Fragmentation Reagents (Ambion, Austin, USA) and then stop with Stop solution. Hybridization was realized using SlideBooster apparatus (Advalytix, Beckman Coulter Biomedical GmbH, Germany).
|
| Scan protocol |
Microarrays were scanned using an Agilent scanner (Agilent Technologies).
|
| Data processing |
Data acquisition was performed using GenePix Pro 6.0 (Axon Molecular Devices, Sunnyvale, CA, USA).
|
| |
|
| Submission date |
Aug 12, 2011 |
| Last update date |
Aug 17, 2011 |
| Contact name |
Svetlana UZBEKOVA |
| E-mail(s) |
[email protected]
|
| Phone |
+33247427951
|
| Organization name |
INRA
|
| Lab |
Physiologie de la Reproduction et des Compartements
|
| Street address |
PRC INRA
|
| City |
Nouzilly |
| ZIP/Postal code |
37380 |
| Country |
France |
| |
|
| Platform ID |
GPL6694 |
| Series (1) |
| GSE31361 |
Gene expression profiling of in vitro matured bovine oocytes cultivated with or without cumulus cells. |
|
