|
| Status |
Public on Jul 10, 2008 |
| Title |
C57 smoked mouse_1 |
| Sample type |
RNA |
| |
|
| Source name |
smoked lung
|
| Organism |
Mus musculus |
| Characteristics |
C57Bl/6J three month-old male mice exposed to the smoke from three cigarettes/day, 5 days/week for 4 weeks
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from mice lungs using Trizol (Invitrogen Life technologies, Irvine, CA) according to the manufacturer’s instructions.
|
| Label |
biotin
|
| Label protocol |
15 µg of RNA were reverse-transcribed into cDNA using SuperScript II kit (Invitrogen Life Technologies, Irvine CA) with an oligo(dT)24 primer containing a T7 polymerase promoter site at 3’ purchased from the MWG Biotech (Ebersberg, Germany). cDNA was purified by phenol-chloroform extraction followed by ethanol precipitation and resuspended in DEPC water. Labeled cRNA was synthesized using the in vitro transcription reaction in the presence of biotin-11-CTP and biotin-16-UTP, and the BioAssay High Yield RNA Transcript Labeling Kit (Enzo Diagnostics, Farmingdale, NY). Biotin-labeled cRNA was cleaned up using the Qiagen RNeasy Mini Kit (Qiagen, Hilden, Germany) and ethanol precipitation.
|
| |
|
| Hybridization protocol |
Hybridization was carried out for 16 hr at 45oC. Chips were then washed with 6x SSPE and stained with streptavidin-phycoerythrin according to Affymetrix’s guidelines.
|
| Scan protocol |
Chips were scanned using a confocal microscope scanner with the 560-nm long-pass filter (HP GeneArray Scanner 2500) according to Affymetrix’s guidelines.
|
| Description |
The mice were housed in groups of two to four in macrolon cages. Room temperature ranged between 22°C and 24°C, the relative humidity ranged between 40% and 50%; food and water were supplied ad libitum.
|
| Data processing |
Expression values were quantified using the statistical algorithms implemented in Affymetrix Micrarray Suite 5.0. Array signals were scaled to an average intensity of 100 and the resulting data were analyzed by GeneSpring Expression Analysis Software (version Gx 7.3).
|
| |
|
| Submission date |
Jul 08, 2008 |
| Last update date |
Jul 09, 2008 |
| Contact name |
Paolo Fardin |
| E-mail(s) |
paolofardin@ospedale-gaslini.ge.it
|
| Organization name |
Istiuto Giannina Gaslini
|
| Department |
Molecular Biology
|
| Street address |
Largo G. Gaslini 5
|
| City |
Genoa |
| ZIP/Postal code |
16147 |
| Country |
Italy |
| |
|
| Platform ID |
GPL81 |
| Series (1) |
| GSE12036 |
mouse lung resistance or sensitivity to cigarette smoke |
|
