|
| Status |
Public on Jan 01, 2015 |
| Title |
Smad3 ChIP, M4 cell, minus, biological rep1 |
| Sample type |
genomic |
| |
|
| Source name |
MCF10CA1a.cl1 ("M4"), vehicle treated
|
| Organism |
Homo sapiens |
| Characteristics |
treatment: vehicle treated
cell line: MCF10Ca1a.cl1 ("M4")
cell line origin: Breast epithelium
tumor stage represented: Malignant, high grade, metastatic
chip antibody: Smad3
chip antibody manufacturer: Abcam
chip antibody catalog #: 28379
|
| Treatment protocol |
Cells were grown to 50-60% confluence and then serum-starved in DMEM/F12 supplemented with Serum Replacement 1 for 16 hours prior to treatment with TGF-beta (5ng/ml: "plus" condition) or vehicle ("minus condition) for 1 hour.
|
| Growth protocol |
M1 and M2 cells were grown in DMEM/F12 medium supplemented with 10ug/ml insulin. 20ng/ml EGF, 0.5ug/ml hydrochortisone, and 100ng/ml cholera toxin and 5% horse serum. M3 and M4 cells were grown in DMEM/F12 medium supplemented with 5% Horse serum.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Chromatin immunoprecipitation was performed using anti-Smad3 antibody (Abcam #28379) and ChIPed DNA was amplified through 2 rounds using a Whole Genome Reamplification kit (Sigma) followed by EtOH precipitation.
|
| Label |
Biotin
|
| Label protocol |
ChIPed DNA was biotinylated according to the standard Affymetrix protocol (Affymetrix Chromatin Immunoprecipitation Assay Protocol (Affymetrix).
|
| |
|
| Hybridization protocol |
Following fragmentation, 10ug biotinylated DNA was hybridized for 16 hr at 45oC on the GeneChip Human Promoter 1.0R Array. GeneChips were washed and stained in the Affymetrix Fluidics Station 450.
|
| Scan protocol |
Genechips were scanned using an Affymetrix GeneChip Scanner 3000 7G and data was collected using Affymetrix AGCC software.
|
| Description |
Smad3 ChIPed DNA
|
| Data processing |
Affymetrix Tiling Analysis Software (TAS, version 1.2.0) was used for data processing. Quantile normalization was performed within each comparison group; quadruplicate for M1 cells and M2 cells or duplicate for M3 cells and M4 cells.
The probe signals and p-values were computed using window size of 200 bp to generate BAR file (TAS Binary Analysis results).
|
| |
|
| Submission date |
Dec 08, 2011 |
| Last update date |
Jan 01, 2015 |
| Contact name |
Howard Yang |
| E-mail(s) |
yanghow@mail.nih.gov
|
| Phone |
2402765257
|
| Organization name |
NCI
|
| Street address |
9609 Medical Center Dr.
|
| City |
Rockville |
| State/province |
MD |
| ZIP/Postal code |
20850 |
| Country |
USA |
| |
|
| Platform ID |
GPL5082 |
| Series (2) |
| GSE34271 |
Smad3 ChIP-chip analysis on human breast epithelial cells of the MCF10A series |
| GSE34277 |
MCF10A-based xenograft model of breast cancer |
|
