|
| Status |
Public on Apr 15, 2009 |
| Title |
453_actein20microg_ml_24hr_rep2 |
| Sample type |
RNA |
| |
|
| Source name |
MDA-MB-453 cell line treated with actein 20 micrograms/ml 24 hours
|
| Organism |
Homo sapiens |
| Characteristics |
MDA-MB-453 human breast cancer cell line
|
| Biomaterial provider |
ATCC
|
| Treatment protocol |
Actein was obtained from ChromaDex (Laguna Hills, CA, lot number 01355-101), purity 89% by HPLC. Actein was dissolved in dimethylsulfoxide (DMSO) (Sigma; St. Louis, MO) prior to addition to the cell cultures. Cells were seeded in 15 cm plates and allowed to attach for 24 h. The medium was replaced with fresh medium containing actein. 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 BioArray high yield RNA transcript labeling kit (Enzo Life Sciences; Farmingdale, NY), according to a modified Affymetrix 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 the BioArray high yield RNA transcript labeling kit (Enzo Life Sciences; Farmingdale, NY), according to a modified Affymetrix 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 20 microgram/ml actein for 24 hours.
|
| Data processing |
Data was normalized using the GCRMA algorithm. Only 24 hour 20microgram/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 09, 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. |
|
