|
| Status |
Public on Apr 15, 2009 |
| Title |
453_dmso_24hr_rep3 |
| Sample type |
RNA |
| |
|
| Source name |
Sigma
|
| Organism |
Homo sapiens |
| Characteristics |
MDA-MB-453 human breast cancer cell line
|
| Biomaterial provider |
ATCC
|
| Treatment protocol |
Dimethylsulfoxide (DMSO) (Sigma; St. Louis, MO) was added to the cell cultures as a control for the effect of the addition of actein. Cells were seeded in 15 cm plates and allowed to attach for 24 h. The medium was replaced with fresh medium containing dmso. The cells were treated for 24 h after which the cells were collected for microarray analysis.
|
| Growth protocol |
Cells were grown in Dulbecco’s Modified Eagle’s medium (DMEM) (Gibco BRL Life Technologies, Inc., Rockville, MD) containing 10% (v/v) fetal bovine serum (FBS) (Gibco BRL) at 37 °C, 5% C02.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total cellular RNA was extracted using Trizol (Invitrogen; Carlsbad, CA) according to the manufacture’s protocol with minor modifications, and then purified twice with Qiagen’s RNeasy column. High quality total RNA (8 μg) was reverse transcribed with T7-(dT)24 primer and Super Script III reverse transcriptase (Invitrogen).
|
| Label |
biotin
|
| Label protocol |
After purification, cDNA was in vitro transcribed into biotin labeled antisense cRNA with the GeneChip® One-Cycle Target Labeling and Control Reagents (Affymetrix, Inc.; Santa Clara, CA), according to the Affymetrix kit protocol.
|
| |
|
| Hybridization protocol |
Total cellular RNA was extracted using Trizol (Invitrogen; Carlsbad, CA) according to the manufacture's protocol with minor modifications, and then purified twice with Qiagen's RNeasy column. High quality total RNA (8 μg) was reverse transcribed with T7-(dT)24 primer and Super Script III reverse transcriptase (Invitrogen). After purification, cDNA was in vitro transcribed into biotin labeled antisense cRNA with with the GeneChip® One-Cycle Target Labeling and Control Reagents (Affymetrix, Inc.; Santa Clara, CA), according to the Affymetrix kit protocol. cRNA (15 μg) was fragmented into the final probe and hybridized to human U 133A 2.0 gene chips (Affymetrix, Inc.; Santa Clara, CA), comprised of more than 22,000 probe sets.
|
| Scan protocol |
Scanning was performed on an Affymetrix 3000-7G Scanner with GCOS software. Scanning was peformed according to the protocol described in the Affymetrix GeneChip® Expression Analysis Technical Manual, November 2004 Edition.
|
| Description |
MDA-MB-453 human breast cancer cell line treated with dmso control for 24 hours.
|
| Data processing |
Data was normalized using the GCRMA algorithm. Only 24 hour 40microgram/ml actein data and the corresponding DMSO controls were included in the normalization (3 treated samples and 3 controls). Differential expression and its statistical significance was obtained using the LIMMA algorithm.
|
| |
|
| Submission date |
May 03, 2007 |
| Last update date |
Apr 15, 2008 |
| Contact name |
Linda Saxe Einbond |
| E-mail(s) |
le2012@columbia.edu
|
| Phone |
(212)305-6924
|
| Organization name |
Columbia University
|
| Department |
Rehabilitation Medicine
|
| Lab |
Einbond
|
| Street address |
701 West 168Th St.
|
| City |
New York |
| State/province |
NY |
| ZIP/Postal code |
10032 |
| Country |
USA |
| |
|
| Platform ID |
GPL571 |
| Series (1) |
| GSE7848 |
Effect of actein on the growth of the MDA-MB-453 breast cancer cell lines as a function of time and concentration. |
|
