|
| Status |
Public on Aug 28, 2020 |
| Title |
RecCFP_Control RecCtrl_rep4 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Inducible MCF7 RecCFP cancer cell line
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: MCF7
treatment: Induced for 48h with doxycycline
phenotype: RecCFP
|
| Growth protocol |
Recombinant RecCtrl, RecCFP and RecCFPm breast cancer cell lines were cultivated under optimal growth conditions and gene expression was induced with 50ng/ml doxycycline for 48 h; a cell pellet was harvested and total RNA was purified according to the extract protocol
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the mirVana™ miRNA Isolation Kit with phenol from Ambion™ (AM1561, Life technologies) following manufacturer's instructions. NanoDrop Spectrophotometer (NanoDrop Technologies) was used for RNA quantification. The quality of extracted RNA was assessed with Bioanalyzer 2100 (Agilent Technologies) using the RNA 6000 Nano Kit (Agilent Technologies).
|
| Label |
Cy3
|
| Label protocol |
1 µg of total RNA were treated with a Spike-In control (5188-5279, Agilent technologies); cDNA synthesis, cRNA synthesis/amplification and cRNA purification was conducted according to the manufacturer`s instructions: Two-Color Microarray-Based Gene Expression Analysis for Low Input Quick Amp Labeling (5190-2306, Agilent technologies)
|
| |
|
| Channel 2 |
| Source name |
Inducible MCF7 RecCtrl cancer cell line
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: MCF7
treatment: Induced for 48h with doxycycline
phenotype: RecCtrl
|
| Growth protocol |
Recombinant RecCtrl, RecCFP and RecCFPm breast cancer cell lines were cultivated under optimal growth conditions and gene expression was induced with 50ng/ml doxycycline for 48 h; a cell pellet was harvested and total RNA was purified according to the extract protocol
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the mirVana™ miRNA Isolation Kit with phenol from Ambion™ (AM1561, Life technologies) following manufacturer's instructions. NanoDrop Spectrophotometer (NanoDrop Technologies) was used for RNA quantification. The quality of extracted RNA was assessed with Bioanalyzer 2100 (Agilent Technologies) using the RNA 6000 Nano Kit (Agilent Technologies).
|
| Label |
Cy5
|
| Label protocol |
1 µg of total RNA were treated with a Spike-In control (5188-5279, Agilent technologies); cDNA synthesis, cRNA synthesis/amplification and cRNA purification was conducted according to the manufacturer`s instructions: Two-Color Microarray-Based Gene Expression Analysis for Low Input Quick Amp Labeling (5190-2306, Agilent technologies)
|
| |
|
| |
| Hybridization protocol |
The hybridization was conducted with the Hybridization Gasket Slide Kit (G2534-60014, Agilent technologies) and the Gene Expression Hybridization Kit (5188-5242, Agilent technologies) following the manufacturer’s instructions and the samples were hybridized on the SurePrint G3 Custom Gene Expression Microarray 8x60K (G4102A, Agilent technologies)
|
| Scan protocol |
SurePrint G3 Custom Gene Expression Microarray, 8x60K (G4102A, Agilent Technologies) was scanned in the Agilent 2505C DNA Microarray Scanner with Sure-Scan High Resolution Technology. Images were quantified using Agilent Feature Extraction Software version 10.7.3.1.
|
| Description |
Biological replicate 4 of 4. Control RecCtrl with empty vector plasmid; RecCFP
|
| Data processing |
Raw data were processed using the limma R package. Background normalization was performed using normexp with an offset of 50. Each slide was normalized using LOESS. Between array quantile normalization was performed last.
Quantile (between arrays) and loess (within arrays) normalized M values.
|
| |
|
| Submission date |
Aug 30, 2017 |
| Last update date |
Aug 28, 2020 |
| Contact name |
Ines Block |
| Organization name |
Odense University Hospital
|
| Department |
Department of Clinical Genetics
|
| Street address |
J.B. Winsløws Vej 4
|
| City |
Odense |
| ZIP/Postal code |
5000 |
| Country |
Denmark |
| |
|
| Platform ID |
GPL13607 |
| Series (1) |
| GSE103302 |
Stable recombinant MCF7 human breast cancer cell lines with inducible expression of CFP (NM_002621.2, RecCFP) or a mutated CFP variant (S27X, RecCFPm) vs. recombinant MCF7 control recombinants expressing an empty expression vector (RecCtrl) |
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