|
| Status |
Public on Apr 05, 2010 |
| Title |
50mM Ethanol-Rep2 |
| Sample type |
RNA |
| |
|
| Source name |
airway epithelial cells, lot number, ethanol treated or vehicle untreated
|
| Organism |
Homo sapiens |
| Characteristics |
human airway epithelial cells were obtained from commercial source
|
| Extracted molecule |
total RNA |
| Extraction protocol |
sample was processed using the Trizol method
|
| Label |
biotin
|
| Label protocol |
5 ug total RNA was used for first strand synthesis using T7-(dT)24 primer (Affymetrix) and SuperScript cDNA Synthesis Kit (Invitrogen). Synthesis of biotinylated-labeled cRNA was carried out using the RNA Transcript Labeling Kit (ENZO).
|
| |
|
| Hybridization protocol |
After the prehybridization of the GeneChip, 15 ug of fragmented cDNA was pipetted into a GeneChip and hybridized overnight in the Hybridization Oven 640 at 60 rpm, washed, stained with streptavidin-phycoerythrin using a microfluidics station as described by the manufacture.
|
| Scan protocol |
Chips were scanned with the High Resolution 3000 Scanner (G7) and signal and background intensities were quantitated by pixel intensity, and expression signals were analyzed using the GeneChip Operating Software (GCOS 1.4). Expresson signals were scaled to a target intensity of 500.
|
| Description |
Cells were grown on millicells inserts of a 24-well tissue dish. Cells were seeded with 10,000 per millicell initially and used at 24,000 to 46,000 cells.
|
| Data processing |
The CEL data file from each array was imported into Genespring GX 7.3 (Agilent Technologies) and preprocessed using the robust multichip average (RMA) method and per gene normalization was applied using the median values of the no EtOH control samples. The CHP files were also imported and the detection call metrics from the CHP files were used to filter out transcripts that were found absent in all nine samples. In addition, transcripts that were not within a standard deviation of 1.4 and exhibiting less than 1.5 fold change were also removed from further analysis. To identify up-regulated and down-regulated transcripts, genes were filtered using the volcano plot with the limit set at p<0.01 and 2.0 fold change. The parametric test, with variance not equal, was applied without multiple test correction.
|
| |
|
| Submission date |
May 07, 2008 |
| Last update date |
Apr 05, 2010 |
| Contact name |
guoshon wang |
| E-mail(s) |
gwang@lsuhsc.edu
|
| Fax |
504-568-8500
|
| Organization name |
LSUHSC
|
| Department |
Gene Therapy
|
| Street address |
533 Bolivar St.
|
| City |
New Orleans |
| State/province |
LA |
| ZIP/Postal code |
70112 |
| Country |
USA |
| |
|
| Platform ID |
GPL570 |
| Series (1) |
| GSE12253 |
Mechanism of biphasic effects of alcohol on gene expression |
|
