|
| Status |
Public on May 29, 2010 |
| Title |
4T1 biological replicate 2 (gene) |
| Sample type |
RNA |
| |
|
| Source name |
4T1
|
| Organism |
Mus musculus |
| Characteristics |
cell line: 4T1
|
| Treatment protocol |
Four mice were injected (1×10^-5 cells) with 168FARN, five mice with 4T07 and four mice with 4T01. Tumor volumes were calculated using the following formula: (?LW^2)/6, where L is the length and W is the width of the tumor. Tumors were surgically removed, using a cautery unit, once they reached a volume between 100 and 125 mm^3.
|
| Growth protocol |
All cell lines were grown in DMEM supplemented with 10% fetal bovine serum, 10 mmol/L HEPES, 1 mmol/L sodium pyruvate, 1.5 g/L sodium bicarbonate, penicillin/streptomycin, and fungizone.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was purified using RNeasy Mini Kit Columns following the manufacturers instructions. We assessed The RNA quality using RNA 6000 NanoChips with the Agilent 2100 Bioanalyzer (Agilent, Palo Alto, USA). Tumors were hybridized independently at the functional genomics facility of McGill University and Genome Quebec Innovation Centre (Montreal, Quebec, Canada).
|
| Label |
Biotin
|
| Label protocol |
Biotin-labelled target for the microarray experiment were prepared using 1µg of total RNA. The RNA was subjected to a rRNA removal procedure with the RiboMinus Human/Mouse Transcriptome Isolation Kit (Invitrogen) and cDNA was synthesized using the GeneChip® WT (Whole Transcript) Sense Target Labeling and Control Reagents kit as described by the manufacturer (Affymetrix)
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| |
|
| Hybridization protocol |
Hybridization was performed using 5 micrograms of biotinylated target, which was incubated with the GeneChip® Human Exon 1.0 ST array (Affymetrix) at 45˚C for 16-20 hours. Following hybridization, non-specifically bound material was removed by washing and detection of specifically bound target was performed using the GeneChip® Hybridization, Wash and Stain kit, and the GeneChip® Fluidics Station 450 (Affymetrix).
|
| Scan protocol |
The arrays were scanned using the GeneChip® Scanner 3000 7G (Affymetrix) and raw data was extracted from the scanned images and analyzed with the Affymetrix Power Tools software package (Affymetrix).
|
| Description |
gene expression array
|
| Data processing |
The Affymetrix Power Tools software package (Affymetrix) was used to quantile normalize the probe fluorescence intensities and to summarize the probe set (representing exon expression) and meta-probe set (representing gene expression) intensities. Transcript assignments were based on mm8 build. The library files for gene expression and exon expression are MoEx-1_0-st-v1.r2.dt1.mm8.core.mps.mask and MoEx-1_0-st-v1.r2.dt1.mm8.full.ps, respectively. Probeset intensities (exon data) : Probe logarithmic intensity error model (PLIER) for probe set intensities. The probeset_dabg.txt file on the Series record: Detection Above BackGround (DABG) statistics for probe set presence/absence. The meta probeset intensities (gene data): Iterative Probe logarithmic intensity error (ITER-PLIER) for meta-probe set intensities.
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| |
|
| Submission date |
May 25, 2010 |
| Last update date |
May 28, 2010 |
| Contact name |
Amandine Bemmo |
| Organization name |
McGill University
|
| Department |
Human Genetics
|
| Lab |
McGill University and Genome Quebec Innovation Center
|
| Street address |
740, Dr Penfield Avenue
|
| City |
Montréal |
| State/province |
Québec |
| ZIP/Postal code |
H3A 1A4 |
| Country |
Canada |
| |
|
| Platform ID |
GPL6096 |
| Series (1) |
| GSE21994 |
Exon-level transcriptome profiling in murine breast cancer reveals splicing changes specific to tumors with different metastatic abilities |
|
| Relations |
| Alternative to |
GSM546975 (exon-level) |
