|
| Status |
Public on Aug 01, 2013 |
| Title |
p53-, rep2 |
| Sample type |
RNA |
| |
|
| Source name |
mammary epithelial cells, p53-
|
| Organism |
Mus musculus |
| Characteristics |
cell type: mammary epithelial cells
gender: female
age: 8 weeks
myc: WT
p53: KO
rb: WT
|
| Treatment protocol |
Adenoviral expression of Cre-recombinase was used to delete the RB and/or p53 genes, and retrovirus was used to achieve MYC overexpression.
|
| Growth protocol |
Mammary epithelial cells were isolated from 8-week-old female mice and sub-cultured in DMEM/F12 media supplemented with 5% horse serum, 20ng/mL EGF, 10μg/ml insulin, 1ng/ml cholera toxin, 100μg/ml hydrocortisone, 100 U/ml penicillin/streptomycin and 2mM L-glutamine at 37oC in 5% CO2.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was harvested from proliferating cell populations using the RNeasy Blood and Tissue kit (Qiagen, Inc., Valencia, CA) according to the manufacturer's suggested protocols, and quantified on a Nanodrop ND-100 spectrophotometer. RNA quality was assessed by analysis on an Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA).
|
| Label |
biotin
|
| Label protocol |
Labeling was performed by the WT-Ovation Pico RNA amplification system (NuGen Technologies, Inc.)
|
| |
|
| Hybridization protocol |
Arrays were hybridized with fragmented and biotin-labeled target (2.5 µg) in 110 µl of hybridization cocktail. Target denaturation was performed at 99oc for 2 min. and then 45oc for 5 min., followed by hybridization for 18 hrs. Arrays then were washed and stained using GeneChip Fluidic Station 450, and hybridization signals were amplified using antibody amplification with goat IgG (Sigma-Aldrich) and anti-streptavidin biotinylated antibody (Vector Laboratories, Burlingame, CA, USA).
|
| Scan protocol |
Arrays were scanned on an Affymetrix GeneChip Scanner 3000, using Command Console Software.
|
| Description |
KM-05
|
| Data processing |
Raw intensity files from the Affymetrix Mouse Gene 1.0 ST GeneChip arrays were processed using Affymetrix Expression Console version 1.1. Gene-level expression measurements were computed using the iterPLIER algorithm on the core probesets and exported with annotation release 32, dated June 23rd, 2011.
|
| |
|
| Submission date |
Aug 31, 2012 |
| Last update date |
Aug 01, 2013 |
| Contact name |
Adam Ertel |
| Organization name |
Thomas Jefferson University
|
| Department |
Cancer Biology
|
| Lab |
BLSB 1009
|
| Street address |
233 S 10TH ST
|
| City |
PHILADELPHIA |
| State/province |
PA |
| ZIP/Postal code |
19107 |
| Country |
USA |
| |
|
| Platform ID |
GPL6246 |
| Series (1) |
| GSE40545 |
RB pathway deregulation promotes invasion and disease progression in a mouse model of MYC-overexpressing mammary tumorigenesis |
|
