|
| Status |
Public on Jun 21, 2014 |
| Title |
MCF-7_siSMRT_24h_E2_rep1 |
| Sample type |
RNA |
| |
|
| Source name |
SMRT-depleted MCF-7_Estradiol treated for 24 h
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: MCF-7
cell type: breast cancer cell line
transfected with: SMRT-targeting siRNA
treated with: 1 nM estradiol (E2) for 24hrs
|
| Treatment protocol |
Cells were plated DME supplemented with 10% FBS 24 h prior to transfection. The transfection was conducted in Opti-MEM using oligofectamine. Media was changed after transfection to phenol red-free DME medium supplemented with 10% charcoal stripped fetal bovine serum and 1% L-Glutamine. After 48 h cells were treated with vehicle (0.1% EtOH) or 1 nM E2 for an additional 4 or 24 h.
|
| Growth protocol |
We routinely grow MCF-7 cells in Dublecco's Modified Eagle Medium (DME) supplemented with 10% fetal bovine serum (FBS).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated using the RNeasy kit from Qiagen.
|
| Label |
biotin
|
| Label protocol |
Fragmentation of single-stranded cDNA used UDG and APE 1 enzymes, this breaks DNA at points at which dUTP was added during the second cycle of reverse transcription. Followed by terminal labeling with terminal deoxyucleotidyl transferase and Affymetrix proprietary DNA Labeling Reagent.
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| |
|
| Hybridization protocol |
Arrays were hybridized for 17 h at 45°C at 60rpm rotation in Affymetrix GeneChip Hybridization Oven 640. Arrays were washed using fluidics script FS450_00070 and stained with streptavidin, R-phycoerythrin conjugate stain using Affymetrix GeneChip Fluidics Station 450. Signal amplification was done using biotinylated anti-streptavidin.
|
| Scan protocol |
Arrays were scanned on Affymetrix GeneChip Scanner 300 and analyzed using Affymetrix GeneChip Operating System.
|
| Description |
SMRT_KD_24h_E2_rep1
|
| Data processing |
Data analysis was performed using GeneChip Operating Software (GCOS) with the RMA alogrithm.
|
| |
|
| Submission date |
May 23, 2014 |
| Last update date |
Jun 21, 2014 |
| Contact name |
Carolyn L Smith |
| E-mail(s) |
carolyns@bcm.edu
|
| Organization name |
Baylor College of Medicine
|
| Department |
Molecular and Cellular Biology
|
| Street address |
One Baylor Plaza
|
| City |
Houston |
| State/province |
Texas |
| ZIP/Postal code |
77030 |
| Country |
USA |
| |
|
| Platform ID |
GPL6244 |
| Series (1) |
| GSE57935 |
Expression data for Control and SMRT-Depleted MCF-7 Cells |
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