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Status |
Public on Apr 30, 2009 |
Title |
DMSO3_24HR |
Sample type |
RNA |
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|
Source name |
DMSO-treated embryonic primary chondrocyte cultures, incubated for 24hrs.
|
Organism |
Mus musculus |
Characteristics |
strain: CD1 tissue: cartilage developmental stage: embryonic 15.5 days
|
Biomaterial provider |
Mouse supplier: Charles River. Cultures were completed in Frank Beier's laboratory by Veronica Ulici and Claudine James and Lee-Anne Stanton
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Treatment protocol |
Primary monolayer chondrocytes were treated with the DMSO control (vehicle) and diluted in fresh media supplemented with 0.25 mM ascorbic acid (Sigma) and 1 mM ß-glycerophosphate (Sigma) and incubated for 24 hrs.
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Growth protocol |
Tibiae, femurs and humeri were isolated from E15.5 mouse embryos and placed in a-mem media (Invitrogen) containing 0.2% bovine serum albumin (BSA), 1 mm ß-glycerophosphate, 0.05 mg/ml ascorbic acid and penicillin/streptomycin incubated at 37°c in a humidified 5% co2 incubator overnight. The following morning media was removed and the bones placed in 4 ml of 0.25% trypsin-edta (invitrogen) for 15 min at 37oc. Trypsin was subsequently replaced with 1 mg/ml collagenase p (roche) in dmem / 10% fetal bovine serum (invitrogen), and cells were incubated at 37°c with rotation at 100 rpm for 90 min. Following digestion, the cell suspension was centrifuged for 5 min at 1000 rpm, and the collagenase containing supernatant was decanted. Chondrocytes were resuspended in media containing 2: 3 DMEM:F12, 10% fetal bovine serum, 0.5 mM L-glutamine, and penicillin/streptomycin (25 units/ml). Cells were seeded in 6-well NUNC plates at a density of 2.5X104 cells per ml and incubated overnight.
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Extracted molecule |
total RNA |
Extraction protocol |
Qiagen RNeasy extraction
|
Label |
biotin
|
Label protocol |
Completed according to the London Regional Genomics Centre (LRGC): http://www.lrgc.ca/
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|
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Hybridization protocol |
LRGC specifications
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Scan protocol |
M.A.S. 5.0-see LRGC specifications
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Description |
DMSO is the vehicle for the pharmacological inhibitors LY294002, TSA, UO126, SB202190 and represents the experimental control.
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Data processing |
MAS 5.0 Algorythm : "The procedure is to construct a plot of each chip's probes against the corresponding probes on the baseline chip; eliminate the highest 1% of probes (and for symmetry the lowest 1%), and fit a regression line to the middle 98% of probes (ie. estimate slope and intercept: two parameters). Transform the values in each probe, by subtracting the intercept, and dividing by the slope, so that the regression line becomes the identity ( y = x ) line." http://discover.nci.nih.gov/microarrayAnalysis/Affymetrix.Preprocessing.jsp
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Submission date |
Mar 02, 2009 |
Last update date |
Apr 21, 2009 |
Contact name |
veronica ulici |
E-mail(s) |
verau79@yahoo.com
|
Phone |
1-519-661-3387
|
URL |
http://www.schulich.uwo.ca/research/skeletal-web/beier/old_news.htm
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Organization name |
University of Western Ontario
|
Department |
Physiology and Pharmacology
|
Lab |
Dr. Frank Beier
|
Street address |
1151 Richmond Street
|
City |
London |
State/province |
Ontario |
ZIP/Postal code |
N6A5C1 |
Country |
Canada |
|
|
Platform ID |
GPL1261 |
Series (1) |
GSE15069 |
Inhibitor trials in chondrocytes - MAS 5.0 normalization |
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