|
| Status |
Public on Jun 30, 2012 |
| Title |
MDA-MB-231 1 μg/ml JAGGED1-FC + 5 μM DAPT 6 hours Treatment Replicate 1 |
| Sample type |
RNA |
| |
|
| Source name |
MDA-MB-231, ER-, mutated p53, basal, breast cancer, 1 μg/ml JAGGED1-FC, 5 μM DAPT, 6 hours
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: MDA-MB-231
estrogen receptor status: Negative
p53 status: Mutant
cancer type: Breast
cancer subtype: Basal
treatment: 1 μg/ml JAGGED1-FC + 5 μM DAPT
duration: 6 hours
|
| Treatment protocol |
For each array sample, 5 million cells were plated either on 1 μg/ml JAGGED1-Fc or 1 μg/ml DLL4-Fc or 1 μg/ml Fc as control with or without 5 μM DAPT or DMSO as the carrier control
|
| Growth protocol |
MCF7 and MDA-MB-231 cells were maintained and expanded in DMEM with 10% FBS and 1% of PEST (Invitrogen) in 5% CO2 at 37°C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from all treated MCF7 and MDA-MB-231 cells after 6 hours ligand stimulation using the RNeasy Mini kit (Qiagen) according to manufacturer instructions. RNA quality was assessed on the 2100 Bioanalyser (Agilent) using the RNA 6000 Nano Chip kit (Agilent) for intact 18S and 28S ribosomal peaks without significant degradation (RNA Integrity Number >9) for all samples.
|
| Label |
Cy3
|
| Label protocol |
500ng of total RNA from each sample was reverse transcribed into cDNA and in vitro transcribed into biotin-labelled cRNA using the Illumina TotalPrep RNA Amplification kit (Ambion).
|
| |
|
| Hybridization protocol |
750ng of each cRNA sample was hybridized to HumanRef-8 v3 Expression BeadChip microarrays (Illumina) as per manufacturer specifications.
|
| Scan protocol |
Microarrays were scanned on the BeadArray Reader (Illumina, USA) at scan factor 1 as per manufacturer specifications.
|
| Description |
Chua SW, Vijayakumar P, Nissom PM, Yam CY, Wong VV, Yang H: A novel normalization method for effective removal of systematic variation in microarray data. Nucleic acids research 2006, 34(5):e38.
|
| Data processing |
Raw intensity values were subjected to background subtraction on the BeadStudio Data Analysis Software (Illumina) and normalized using the cross-correlation method (Chua et al., 2006). Differential gene expression was identified based on a fold change cutoff of >1.5 compared to the average of the Fc controls for each cell line.
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| |
|
| Submission date |
Feb 23, 2012 |
| Last update date |
Jun 30, 2012 |
| Contact name |
Kian Leong LEE |
| E-mail(s) |
kianleong.lee@duke-nus.edu.sg
|
| Phone |
+(65) 6601 3685
|
| Organization name |
National University of Singapore (NUS)
|
| Department |
Duke-NUS Medical School
|
| Lab |
Cancer & Stem Cell Biology Program (CSCB)
|
| Street address |
#07-21, 8 College Road
|
| City |
Singapore |
| State/province |
Singapore |
| ZIP/Postal code |
169857 |
| Country |
Singapore |
| |
|
| Platform ID |
GPL6883 |
| Series (1) |
| GSE36051 |
Non-canonical Notch signaling activates IL-6/JAK/STAT signaling in breast tumor cells and is controlled by p53 and IKKβ |
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