|
| Status |
Public on Dec 31, 2006 |
| Title |
17 A Rapa 0.01/Vehicle/6h/NG |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
RNA prepared from 1 day old whole rat pups
|
| Organism |
Rattus norvegicus |
| Characteristics |
1 day old whole rat pups
|
| Biomaterial provider |
Rat aortic smooth muscle cells were generated in the Cam Patterson lab
|
| Treatment protocol |
Untreated rat pups for reference RNA
|
| Growth protocol |
none
|
| Extracted molecule |
total RNA |
| Extraction protocol |
GITC/Phenol/CHcl3 extraction CsCl2 banding
|
| Label |
Cyanine-3
|
| Label protocol |
Starting with 0.5 ug of total RNA, Cyanine-5 labeled cRNA was produced using the Agilent Low-Input RNA Amplification labeling kit according to manufacturer’s protocol.
|
| |
|
| Channel 2 |
| Source name |
Rat aortic smooth muscle (tmt: Rapa 0.01/Vehicle/6h/NG)
|
| Organism |
Rattus norvegicus |
| Characteristics |
Cell line: Rat aortic smooth muscle cells (RASMC)
|
| Biomaterial provider |
Rat aortic smooth muscle cells were generated in the Cam Patterson lab
|
| Treatment protocol |
Cells were grown in 100 mm dishes in 10% FBS/DMEM to approximately 70-80 % confluence, and were washed once in serum-free DMEM, and then incubated for 16 h in DMEM containing 0.5 % (v/v) FBS. The cells were then stimulated with drug/vehicle, insulin/vehicle in NG/HG for 6/24 hours and were extracted for RNA.
|
| Growth protocol |
The rat aortic smooth muscle cells were maintained at 37oC (5% CO2) in Dulbecco Minimal Essential Medium supplemented with 10% FBS, 2 mM L-glutamine, 100U/ml penicillin and 100 mg/ml streptomycin.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
The cell monolayer was rinsed twice in cold PBS, 5 ml each. Each plate of cells was then lysed in 0.6 ml of Buffer RLT (RNeasy Mini Kit, Qiagen) and total cellular RNA was prepared following the RNeasy protocol.
|
| Label |
Cyanine-5
|
| Label protocol |
Starting with 0.5 ug of total RNA, Cyanine-5 labeled cRNA was produced using the Agilent Low-Input RNA Amplification labeling kit according to manufacturer’s protocol.
|
| |
|
| |
| Hybridization protocol |
2ug sample and 2ug reference amplified cRNAs were fragmented and hybridized to the array for 16 hours in a rotating hybridization oven using the Agilent Target Hybridization Controls and protocol.
|
| Scan protocol |
Slides were washed as indicated in the Agilent protocol. Arrays were then scanned with an Axon GenePix 4000b. Data was obtained using the GenePix Pro 6 software package.
|
| Description |
Gene expression analysis was conducted using Agilent G4130A Rat 22K microarray slides. Starting with 0.5 ug of total RNA, Cyanine-5 labeled cRNA was produced using the Agilent Low-Input RNA Amplification labeling kit according to manufacturer’s protocol. 2ug sample and 2ug reference amplified cRNAs were fragmented and hybridized to the array for 16 hours in a rotating hybridization oven using the Agilent Target Hybridization Controls and protocol. Slides were washed as indicated in the Agilent protocol. Arrays were then scanned with an Axon GenePix 4000b. Data was obtained using the GenePix Pro 6 software package.
|
| Data processing |
loess normalization, log2 transformation
|
| |
|
| Submission date |
Jul 18, 2006 |
| Last update date |
Jul 28, 2006 |
| Contact name |
Monte S Willis |
| E-mail(s) |
[email protected]
|
| Phone |
919-843-1938
|
| Fax |
919-843-4585
|
| Organization name |
UNC
|
| Department |
Pathology & Laboratory Medicine
|
| Lab |
CCBC
|
| Street address |
103 Mason Farm Road
|
| City |
Chapel Hill |
| State/province |
NC |
| ZIP/Postal code |
27759-7525 |
| Country |
USA |
| |
|
| Platform ID |
GPL890 |
| Series (1) |
| GSE5337 |
Gene Expression Profiling In Rat Smooth Muscle Cells Modulated by Rapamycin and Paclitaxel. |
|
