|
| Status |
Public on May 25, 2012 |
| Title |
Wild type day 3 O2 replicate 3 |
| Sample type |
RNA |
| |
|
| Source name |
Homogenized lung tissue
|
| Organism |
Mus musculus |
| Characteristics |
genotype variation: Nrf+/+
treatment: hyperoxia
developmental stage: neonate
|
| Treatment protocol |
Neonatal mice and their foster dams were exposed to room-air exposure. Food and water were provided ad libitum. In 24 hr after birth (P1, saccular stage of lung maturation), neonatal mice were placed in cages in a hyperoxia chamber, and were exposed to 100 percent (≥95 percent) O2 (UHP grade, Min. purity 99.994 percent O2 tanks, National Welders, Durham, NC) 23 hr/day continuously for 1, 2, or 3 d with their foster dams. Water and feed were provided ad libitum during the exposure. Neonatal mice and their foster dams assigned to room-air exposure were placed in cages placed on counter top with food and water provided ad libitum for the same exposure duration. The temperature (72±3 degree F) and humidity (50±15 percent) of the chamber were monitored, and mice were exposed to a 12-hr light-dark cycle. The hyperoxia chamber was opened once each morning for 1 hr to replace foster dams and to check animal health (morbidity and mortality) and water and feed.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from lung homogenates of each mouse (n=3 per group) using the Qiagen RNeasy Mini kit (Qiagen, Valencia, CA) and then evaluated by an Agilent 2100 Bioanalyzer analysis (Agilent Technologies, Palo Alto, CA).
|
| Label |
biotin
|
| Label protocol |
cDNA was synthesized from RNA using a one-cycle cDNAsynthesis kit. Biotinylated RNA was then synthesized from cDNA using an IVT labeling kit (Affymetrix 3'IVT Expression) after incubating at 37◦C for 16 hr.
|
| |
|
| Hybridization protocol |
The biotinylated RNA (12.5 ug) was fragmented and 10 ug was hybridized to the Affymetrix array for 16 hr at 45◦C in a hybridization oven rotating at 60 rpm using the Affymetrix Eukaryotic Target Hybridization Controls and protocol. The array was stained with streptavidin/phycoerythrin in an Affymetrix Fluidics Station 450 using a double-antibody staining procedure and then washed for antibody amplification according to the GeneChip Hybridization, Wash and Stain Kit and user manual.
|
| Scan protocol |
The array was scanned in an Affymetrix Scanner 3000.
|
| Description |
Comparisons: Hyperoxia effects on wild type/3 day treatment, Genotype effects on daily hyperoxia/3 day treatment
|
| Data processing |
The chip data was obtained using the GeneChip Command Console Software (AGCC, Version 1.1) and analyzed using Affymetrix GeneChip Operating Software (GCOS) version 1.2. The CEL files for hybridizations were log2-transformed by use of RMA Express (Bioinformatics 2003 19(2):185-193 and Biostatistics 2003 4(2):249-264).The expression value (average difference) for each gene was determined by calculating the average of differences in intensity (perfect match intensity minus mismatch intensity) between its probe pairs.
|
| |
|
| Submission date |
May 31, 2011 |
| Last update date |
May 25, 2012 |
| Contact name |
Steven R Kleeberger |
| E-mail(s) |
kleeber1@niehs.nih.gov
|
| Organization name |
National Institute of Environmental Health Sciences, NIH
|
| Lab |
Laboratory of Respiratory Biology
|
| Street address |
111 TW Alexander Dr., Building 101, MD D-201
|
| City |
Research Triangle Park |
| State/province |
NC |
| ZIP/Postal code |
27709 |
| Country |
USA |
| |
|
| Platform ID |
GPL1261 |
| Series (1) |
| GSE29632 |
Effect of Nrf2 deletion in postnatal lung development and BPD phenotype in newborn mice |
|
