|
| Status |
Public on Dec 31, 2016 |
| Title |
SP1, biological rep2 |
| Sample type |
RNA |
| |
|
| Source name |
SCC172, SP1 fraction
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: SCC172
fraction: SP1
cell type: oral squamous cell carcinoma (OSCC) cells
|
| Treatment protocol |
Side population analysis was based on the previously described method (Goodell et al., 1997) with slight modifications. Briefly, cells were incubated (1x10^6cells/ml) in pre-warmed DMEM supplemented with 2% fetal bovine serum (FBS) (Sigma) and 10Mm HEPES (Sigma) containing 5 µg/ml Hoechst 33342 for 45 minutes at 37C (water-bath) with intermittent mixing. After incubation, stained cells were washed and re-suspended in ice-cold HBSS (Sigma) supplemented with 2% FBS. Propidium iodide (1µg/ml) was added immediately before analysis to label dead cells. Analysis and sorting was performed on FACS Aria II (BD Biosciences) using Diva Software.
|
| Growth protocol |
The OSCC cell line SCC172 were adherent cells maintained in Dulbecco’s Modified Eagle’s Medium (Sigma, D1152) containing 25mM HEPES and 3.6g/L sodium bicarbonate supplemented with 10% Foetal Bovine Serum (FBS, Sigma) and 1% antibiotic antimycotic cocktail (Invitrogen). Cells were incubated at 37C with 5% CO2 in a humidified incubator.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using Qiagen’s All Prep DNA/RNA Mini Kit (Cat. No. 80204) following the manufacturer’s instructions.
|
| Label |
Biotin
|
| Label protocol |
100ng of total RNA were used for preparation of biotin-labeled targets (cRNA) using modified MessageAmp™ -based protocols (Ambion Inc., Austin, TX).
|
| |
|
| Hybridization protocol |
Labeled cRNA was fragmented in a 0.5ug/uL reaction and used for array hybridization and washing according to the standard Affymetrix protocol on an Affymetrix FS450 Fluidics station.
|
| Scan protocol |
The arrays were scanned on an Affymetrix GeneChip Scanner 3000 7G using the Affymetrix Statistical Algorithm MAS 5.0 (GCOS v1.3) algorithm.
|
| Description |
Differential expression analysis of genes in SP1.
|
| Data processing |
Softwares used for analysis were Partek Genomic Solutions 6.2 (Partek Inc., St. Charles, MO), Matlab 7 (Mathworks Inc., MA) and GCOS 1.3 (Affymetrix Inc., CA).
|
| |
|
| Submission date |
Feb 27, 2012 |
| Last update date |
Dec 31, 2016 |
| Contact name |
Vinitha Richard |
| E-mail(s) |
[email protected]
|
| Phone |
+91 9972844405
|
| Organization name |
Rajiv Gandhi Centre for Biotechnology
|
| Department |
Cancer Research Program
|
| Lab |
MRP Lab
|
| Street address |
Thycaud P.O.
|
| City |
Thiruvananthapuram |
| State/province |
Kerala |
| ZIP/Postal code |
695014 |
| Country |
India |
| |
|
| Platform ID |
GPL571 |
| Series (1) |
| GSE36111 |
Expression data from side population cells of the human OSCC cell line SCC172 |
|
