|
| Status |
Public on Dec 22, 2008 |
| Title |
Lung_C57BL_21 days unexposed to P. murina_rep1 |
| Sample type |
RNA |
| |
|
| Source name |
Mouse lung homogenate
|
| Organism |
Mus musculus |
| Characteristics |
Strain: C57BL, Gender: female, Age: 8 weeks
|
| Growth protocol |
Animals were cohoused with a seeder mouse highly infected with Pneumocystis murina and sacrificed at the corresponding day of exposure, and lungs were harvested at that time. Unexposed animals (controls) were sacrificed at the corresponding time points and lungs were harvested.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using RNeasy Mini Kit and Qiashredder column.
|
| Label |
Biotin
|
| Label protocol |
The cDNA was purified following the procedures described by Affymetrix and was transcribed in vitro and biotin-labeled using the BioArrayHigh Yield RNA Transcript Labeling Kit (ENZO Life Science Inc., Farmingdale, NY).
|
| |
|
| Hybridization protocol |
Twenty mg of fragmented cRNA was used to prepare the hybridization cocktail as described by Affymetrix. Hybridization cocktail containing 15 µg of fragmented cRNA were added to an MG-U74Av2 array (Affymetrix) and hybridized for 16 hours at 45˚C in a Hybridization Oven 640 (Affymetrix). The arrays were stained and washed using the GeneChip® Fluidics Station 400 using R-Phycoerythrin Streptavidin (Molecular Probes) and a biotinylated goat anti-Streptavidin antibody (Vector Laboratories, Burlingame, CA).
|
| Scan protocol |
Experiments 1 and 2 were scanned using the GeneArray® 2500 scanner. The GeneChip® Scanner 3000 was used to scan Experiments 3 and 4. The signal intensity was quantitated using GeneChip® Operating Software v1.2 (GCOS 1.2)
|
| Description |
SG-U21-1
|
| Data processing |
Data were first normalized to the median value of each chip, then logarithmically transformed after the addition of a small value (1% of the median) to dampen the influence of extremely small data values. A one-way ANOVA was performed to determine the significance of changes within each experiment. Affymetrix genes were selected if the P-value was less than 0.001 and the fold change (FC) between the exposed compared to the unexposed animals was at least 1.5 fold or at most 1/1.5 fold .
|
| |
|
| Submission date |
Mar 31, 2008 |
| Last update date |
Dec 22, 2008 |
| Contact name |
Joseph A. Kovacs |
| E-mail(s) |
jkovacs@mail.nih.gov
|
| Organization name |
NIH
|
| Department |
CCMD
|
| Street address |
Bldg. 10, Room 2C145, MSC1662
|
| City |
Bethesda |
| State/province |
MD |
| ZIP/Postal code |
20892 |
| Country |
USA |
| |
|
| Platform ID |
GPL81 |
| Series (1) |
| GSE11005 |
Immune Responses to Pneumocystis Infection are Robust in Immunocompetent Mice but Absent in CD40 Ligand Deficient Mice |
|
