|
| Status |
Public on May 20, 2011 |
| Title |
MCF7_untreated_rep2 |
| Sample type |
RNA |
| |
|
| Source name |
MCF7 breast carcinoma cells, untreated
|
| Organism |
Homo sapiens |
| Characteristics |
MCF7 human breast carcinoma cells, stably transfected with the pTet-Off-regulator plasmid and the pTRE2hyg response plasmid (Clontech).
Cell clone 2.
|
| Biomaterial provider |
A clone of MCF7 cells stably expressing the pTet-Off-regulator plasmid was kindly provided by Dr. Sam W. Lee (Massachusetts General Hospital, Boston, MA).
|
| Treatment protocol |
Cells were grown for 12 days in their normal growth medium.
|
| Growth protocol |
Cells were cultured in DMEM, containing 10% fetal bovine serum, 100 U/ml penicillin, 100 μg/ml streptomycin and selection antibiotics (150 µg/ml G418 and 60 µg/ml hygromycin B) for 12 days prior to extraction of total RNA.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Approx. 60% confluent cells in a 10cm-cell culture dish were washed twice with cold PBS, before total cellular RNA was extracted using 1 ml Trizol reagent (Invitrogen), according to the manufacturer's instructions. The air-dried RNA pellet was dissolved in 50 µl RNAse/DNAse-free water.
|
| Label |
Digoxigenin-UTP
|
| Label protocol |
Digoxigenin-UTP labeled cRNA was generated from 1 µg of total RNA for each sample using Applied Biosystems NanoAmp RT-IVT Labeling Kit (PN 4365715) according to the manufacturer’s protocol.
|
| |
|
| Hybridization protocol |
Array hybridization, array processing, chemiluminescence detection, image acquisition, and analysis were performed using Applied Biosystems Chemiluminescence Detection Kit (PN 4342142) and Applied Biosystems 1700 Chemiluminescent Microarray Analyzer according to the manufacturer’s protocol.
|
| Scan protocol |
Scanning was performed according to the Chemiluminescent Detection Kit (PN 4339627) and Chemiluminescent Microarray Analyzer User Guide (PN 4338852).
|
| Description |
Replicate 2 of 2.
|
| Data processing |
The Expression Array System Software suite performs the auto-gridding, feature extraction, fluorescence normalization, and signal data generation. The quantification data contain many distinct measurements for each probe, including the three basic measurements: Signal, S/N, and Flag values. The Signal value is the fully corrected, background subtracted measurement of chemiluminescent signal for gene expression values. The S/N value represents the ratio of signal above noise, or the measurement uncertainty of the probe signal, and can be used as a confidence level for the probe measurement. An S/N of 3 represents approximately a 99.95% confidence that the probe is detected above the background noise. In situations where the probe showed S/N < 1, the signal measurement is replaced with a 1 SDEV upper limit based on its probe signal SDEV.
Quantile normalization of the raw signals was performed using R software version 2.3.1 (The R Foundation for Statistical Computing).
|
| |
|
| Submission date |
Dec 12, 2008 |
| Last update date |
May 20, 2011 |
| Contact name |
Leah N Cueni |
| E-mail(s) |
leah.cueni@bluewin.ch
|
| Organization name |
ETH Zurich
|
| Department |
Institute of Pharmaceutical Sciences
|
| Street address |
Wolfgang Pauli-Str. 10
|
| City |
Zurich |
| ZIP/Postal code |
8093 |
| Country |
Switzerland |
| |
|
| Platform ID |
GPL2986 |
| Series (1) |
| GSE13926 |
MCF7 tet-off cells treated or not with 80 ng/ml doxycycline for 12d |
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