|
| Status |
Public on Mar 15, 2016 |
| Title |
Control Casodex 3 (48h) |
| Sample type |
RNA |
| |
|
| Source name |
Control siRNA, 1μM casodex 48 hours
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: LNCaP
cell type: prostate cancer
|
| Treatment protocol |
2x10^6 LNCaP cells were electroporated with 800 pmoles of noncoding control siRNA or NCOR1 specific smart pool siRNA (Thermo Fisher Scientific) using R Buffer (Lonza) and Nucleofector Device (Lonza). Cells were plated in 10 cm dishes in RPMI + 10%FBS medium. 24 hours later cells were treated with 1μM casodex or ethanol vehicle. Cells were harvested 48 hours after treatment.
|
| Growth protocol |
LNCaP cells were grown in RPMI medium supplemented with 10%FBS and treated as described with either ethanol vehicle or 1μM casodex for 48 hours
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was prepared with Trizol (Life Technologies) as recommended by the manufacturer and additional reprecipitations with isopropanol and ethanol were performed.
|
| Label |
Biotin
|
| Label protocol |
Samples were labeled using the standard Affymetrix T7 oligo(dT) primer protocol (GeneChip Expression Analysis Technical Manual, Rev 2, Eukaryotic One-Cycle target preparation). Five ug of total RNA was reverse transcribed to produce double-stranded cDNA. The cDNA product was used as a template for the vitro transcription reaction, producing biotin-labeled cRNA. The labeled cRNA was qualified using the Agilent Bioanalyer.
|
| |
|
| Hybridization protocol |
15.0 μg of the labeled cRNA was fragmented and re-checked for concentration. A hybridization cocktail containing Affymetrix spike-in controls and fragmented labeled cRNA was loaded onto a GeneChip® array. The array was hybridized overnight at 45°C with rotation at 60 rpm then washed and stained with a streptavidin, R-phycoerythrin conjugate stain. Signal amplification was done using biotinylated antistreptavidin.
|
| Scan protocol |
The stained array was scanned on the Affymetrix GeneChip® Scanner 3000.
|
| Data processing |
The images were analyzed and quality control metrics recorded using Affymetrix GCOS software version 1.1.2, standard settings.
|
| |
|
| Submission date |
Aug 25, 2014 |
| Last update date |
Mar 15, 2016 |
| Contact name |
Leif E Peterson |
| E-mail(s) |
LEPETERSON@HOUSTONMETHODIST.ORG
|
| Phone |
(713)441-6121
|
| Organization name |
HMRI
|
| Department |
Center for Biostatistics
|
| Street address |
6565 Fannin, Suite MGJ6-031
|
| City |
Houston |
| State/province |
TX |
| ZIP/Postal code |
77030 |
| Country |
USA |
| |
|
| Platform ID |
GPL570 |
| Series (1) |
| GSE60722 |
Regulation of casodex-dependent AR activity by NCOR1 |
|
