|
| Status |
Public on Oct 19, 2011 |
| Title |
B6.IL10 24 ozone 1 |
| Sample type |
RNA |
| |
|
| Source name |
KO_24_O3_1c
|
| Organism |
Mus musculus |
| Characteristics |
strain: B6.129P2-Il10tmlCgn/J
genotype variation: Il10-/-
age: 6-8 weeks
Sex: Male
tissue: Left lung
supplier: Jackson Labs, Bar Harbor, ME
exposure time: 24 hours
agent: ozone
exposure dose: 0.3 ppm O3
test article cas no: 10028-15-6
|
| Treatment protocol |
Mice were placed in individual stainless-steel wire cages within a Hazelton 1000 chamber (Lab Products, Maywood, NJ) equipped with a charcoal and high-efficiency particulate air–filtered air supply. Mice were exposed to 0.3 ppm O3 or filtered air for 24, 48 or 72 hr (23.5 hr/day) as described previously (Cho et al, 2007).
|
| Growth protocol |
We provided mice with water and pelleted open-formula rodent diet NIH-31 (Zeigler Brothers, Gardners, PA) ad libitum. All experimental procedures were conducted in accordance with approved National Institutes of Health Humane Care and Use of Laboratory Animals guidelines and the American Physiological Society’s “Guiding Principles in the Care and Use of Animals.” Animals were treated humanely and with regard for alleviation of suffering.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Left lobes of mouse lung tissue were harvested, flash frozen in liquid nitrogen, and stored at -80oC. For RNA extraction, the lung tissue was homogenized in 2ml Trizol (Invitrogen, Calsbad, CA) with a Tekmar Tissumizer and saw-tooth generator. One ml of homogenate was subsequently processed according to the manufacturer’s (Invitrogen) protocol with the following minor modifications. Two µl of 5 mg/ml glycogen was used as a carrier for the isopropanol precipitation, duration of the isopropanol precipitation was increased to overnight, and all centrifugation times were increased to 15 minutes. RNA pellets were resuspended in nuclease-free water. RNA quality was tested using a Beckman DU680 spectrophotometer, and quality assessment was determined by RNA Nano LabChip analysis on an Agilent Bioanalyzer 2100. A Qiagen RNeasy total RNA cleanup protocol was subsequently performed followed by re-quantitation by spectophotometry.
|
| Label |
Biotin
|
| Label protocol |
Double stranded cDNA was synthesized from 4µg of total RNA using the GeneChip Expression 3’ amplification reagents one-cycle cDNA synthesis kit (Affymetrix). The resultant double-stranded cDNA was column-purified using the GeneChip Sample Cleanup Module. Biotinylated cRNA was synthesized from the double-stranded cDNA by in vitro transcription (IVT) using the GeneChip Expression 3’ amplification reagents for IVT labeling (Affymetrix), and was subsequently purified by column purification with the GeneChip Sample Cleanup Module (Affymetrix), and quantified. Quality of biotinylated cRNAs was assessed by RNA Nano LabChip analysis on an Agilent Bioanalyzer 2100.
|
| |
|
| Hybridization protocol |
A fragmentation buffer (250mM Tris acetate pH 8.1, 150 mM MgOAc, 500mM KOAc) was used to fragment 15µg of cRNA. The fragmented cRNA was then hybridized to GeneChips (Affymetrix Mouse Genome 430A 2.0).
|
| Scan protocol |
Scanned on an Affymetrix scanner.
|
| Description |
Gene expression data from left lung samples from B6.129P2-Il10tmlCgn/J male mice exposed to 0.3 ppm O3 for 24 hours
|
| Data processing |
RMA Express, Visual data mining and statistical analysis
|
| |
|
| Submission date |
Nov 03, 2010 |
| Last update date |
Oct 19, 2011 |
| Contact name |
Steven R Kleeberger |
| E-mail(s) |
[email protected]
|
| 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 |
GPL8321 |
| Series (1) |
| GSE25095 |
Protective Role of IL-10 in Ozone-induced Pulmonary Inflammation |
|
