|
| Status |
Public on Jul 04, 2011 |
| Title |
B7 antimir-30 |
| Sample type |
RNA |
| |
|
| Source name |
hMADS cells, B7, antimir-30
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: human adipose tissue-derived stem cells
clone: B7
agent: antimiR-30
|
| Treatment protocol |
All transfections were performed with HighPerfect transfection reagent (Qiagen). Adipogenic medium was composed of DMEM/Ham’s F12 media supplemented with 10 µg/ml transferrin, 0.86 µM insulin, 0.2 nM triiodothyronine, 1 µM dexamethasone, 100 µM isobutyl-methylxanthine and 1 µM rosiglitazone. Three days later, the medium was changed (dexamethasone and isobutyl-methylxanthine were omitted).
|
| Growth protocol |
Cells were maintained in the proliferation medium, which is composed of DMEM (low glucose) containing 10% fetal calf serum (FCS), 0.01 M HEPES, 100 U/ml penicillin and streptomycin.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted with the Qiagen Rneasy kit. RNA was quantified using a NanoDrop-1000 spectrophotometer and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent).
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 200 ng RNA using the Low Input Quick Amp Labeling Kit, one-color (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
|
| |
|
| Hybridization protocol |
600 ng of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/µg cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 250 ml containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturer's instructions. On completion of the fragmentation reaction, 250 ml of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent SurePrint G3 Human GE 8x60K for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent).
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2565) using one color scan setting for 8x60k array slides (Scan Area 61x21.6 mm, Scan resolution 3µm, Dye channel is set to Green and Green PMT is set to 100%).
|
| Description |
Gene expression after 4 days in adipogenic medium.
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 10.7.3.1 (Agilent) using default parameters (protocol GE1_107_Sep09 and Grid: 028004_D_20101102) to obtain background subtracted and spatially detrended Processed Signal intensities.
|
| |
|
| Submission date |
May 17, 2011 |
| Last update date |
Jul 04, 2011 |
| Contact name |
Kevin Lebrigand |
| Organization name |
IPMC/CNRS
|
| Lab |
Functional Genomics Platform of Nice-Sophia-Antipolis, France.
|
| Street address |
660 route des lucioles
|
| City |
Valbonne - Sophia-Antipolis |
| ZIP/Postal code |
06560 |
| Country |
France |
| |
|
| Platform ID |
GPL13607 |
| Series (1) |
| GSE29207 |
Role of miR-30 miRNAs during adipogenesis |
|
