|
| Status |
Public on Jun 27, 2013 |
| Title |
caffeine- 2 |
| Sample type |
RNA |
| |
|
| Source name |
granulocytes not treated with caffeine
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6J
cell type: granulocytes
|
| Treatment protocol |
Cells were plated at a density of 3x105 cells per ml in two different tissue culture plates. The optimal concentration of caffeine (15 mM) was added to the culture medium for 4 hours before cells were washed twice with PBS. Actinomycin D (2 μg/ml) together with caffeine was then added into one plate whereas only actinomycin D was added to the other to block transcription of genes. After an additional 4 hours incubation, total RNA was collected and used for microarray analysis.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Trizol extraction of total RNA was performed according to the manufacturer's instructions.
|
| Label |
Biotin
|
| Label protocol |
Biotinylated cDNA were prepared according to the Encore Biotin Module (Nugen) from 300 ng total RNA.
|
| |
|
| Hybridization protocol |
Following fragmentation, samples were injected into Affymetrix array chips and hybridized at 45°C and 60 rpm for 17 hours in a hybridization oven (Affymetrix). GeneChips were stained and washed in the Affymetrix GeneChip Fluidic Station 450
|
| Scan protocol |
GeneChips were scanned using Affymetrix GeneChip Scanner 3000 7G
|
| Data processing |
The data was analysed using R with the RUV procedure described in http://statistics.berkeley.edu/tech-reports/800 for scaling and normalization.
|
| |
|
| Submission date |
Jun 26, 2013 |
| Last update date |
Jun 27, 2013 |
| Contact name |
william ritchie |
| Organization name |
centenary institute
|
| Street address |
Building 93
|
| City |
Camperdown |
| State/province |
NSW |
| ZIP/Postal code |
2050 |
| Country |
Australia |
| |
|
| Platform ID |
GPL6246 |
| Series (2) |
| GSE48306 |
Orchestrated intron retention regulates normal granulocyte differentiation [Affymetrix arrays] |
| GSE48307 |
Orchestrated intron retention regulates normal granulocyte differentiation |
|
