|
| Status |
Public on Jul 12, 2008 |
| Title |
C-20314-T/P(4hSTI571)_U74C |
| Sample type |
RNA |
| |
|
| Source name |
TEL-PDGF-betaR(4h, inhibitor STI571)
|
| Organism |
Mus musculus |
| Characteristics |
32Dcl3-TEL-PDGF-betaR
|
| Treatment protocol |
To completely inhibit kinase activity in a short period of time (4 hours), a concentration of inhibitor that was at least 10 times higher than IC50 for its respective activated tyrosine kinase was used (5 uM for imatinib, 0.5 uM for MLN518
|
| Growth protocol |
Cells were maintained in RPMI 1640 with 10% FBS and 1mg/ml G418
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from cells using RNAeasy Mini Kit (Qiagen, CA). Total RNA was reverse transcribed with Superscript Choice System cDNA synthesis kit (Invitrogen, CA) in the presence of an oligo dT-T7 primer (GENSET, CA). After phenol/chloroform extraction and ethanol precipitation, the cDNA pellet was air dried briefly and resuspended in 12 ul of RNAase-free water.
|
| Label |
biotin
|
| Label protocol |
The cDNA was used for in vitro transcription with the BioArray Highyield RNA Transcript Labeling Kit (ENZO, NY).
|
| |
|
| Hybridization protocol |
After cleaning up with Rneasy Mini Kit, labeled cRNA was fragmented by incubation at 95 oC for 35 minutes in fragmentation buffer (40mM Tris-acetate, pH8.1, 100mM KOAc and 30mM MgOAc). The fragmented cRNA was hybridized against Affymetrix Murine Genome U74v2 set oligonucleotide arrays.
|
| Scan protocol |
GeneChips were scanned using GeneArray Scanner with Affymetrix Microarray Suite version 5
|
| Description |
Gene expression data
|
| Data processing |
The arrays were analyzed using GeneCluster 2.0 software (Whitehead/MIT Center for Genome Research, Cambridge, MA).
Data was preprocessed so that the absolute values were between 20 and 16000. A variation filter was applied to the data so that the ratio of maximum value to minimum value was greater than 3.0, and the difference between maximum value and minimum value was greater than 100 in the same row. The filtered datasets were then analyzed by neighborhood analysis to identify genes of interest.
|
| |
|
| Submission date |
Jun 20, 2008 |
| Last update date |
May 26, 2011 |
| Contact name |
Winnie F Tam |
| E-mail(s) |
[email protected]
|
| Phone |
617-355-9079
|
| Fax |
617-355-9093
|
| Organization name |
Brigham and Women's Hospital, Harvard Medical School
|
| Department |
Hematology
|
| Lab |
Dr. Gary Gilliland
|
| Street address |
1 Blackfan Circle
|
| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02453 |
| Country |
USA |
| |
|
| Platform ID |
GPL83 |
| Series (1) |
| GSE11794 |
Untreated 32Dcl3 cell lines expressing oncogenic tyrosine kinases or cells treated with small molecule inhibitors |
|
