|
| Status |
Public on May 05, 2015 |
| Title |
HeLa_Treated_rep2 |
| Sample type |
RNA |
| |
|
| Source name |
HeLa_1 uM arsenite (iAs-T) for 36 d
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: HeLa
treated with: 1 uM of sodium arsenite (iAs-T) for 36 days
|
| Treatment protocol |
HeLa cells were continuously exposed to Vehicle Control (deionized water) or 1 uM of arsenite (Na3AsO3, Sigma-Aldrich), respectively. When reaching about 80-90% confluence, cells were subcultured and Na3AsO3 was then added to cells after overnight attachment. These procedures were repeated every 3 or 4 days for 16 weeks. During the exposure period, cell morphology changes were monitored. Cell malignant transformation was assessed by changes in cell morphology and EMT markers protein levels.
|
| Growth protocol |
Cell cultures were incubated at 37o C with humidified air and 5% CO2. Cells were harvested after each culture ensuring that the cells had ~95% cell viability. Harvested cells were centrifuged for five minutes at 200 x g. Samples were taken each day for counting. Cell viability was determined by trypan blue dye exclusion using a hemacytometer and calculated as percent viability times total cells/ml.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated from cells using an miRNeasy mini kit (Qiagen) and quality assessment was conducted using RNA 6000 Nano-labchip (Bioanalyzer, Agilent) and by a Nanodrop spectrophotometer (Thermo). Total RNA RIN values were between 7.7 and 9 (average 8.65).
|
| Label |
biotin
|
| Label protocol |
Total RNA (100ng) was processed in parallel with an external MAQC A RNA to control robustness of data. Labeled DNA mean yield was min: 13.6 ug; max: 16.7 ug).
|
| |
|
| Hybridization protocol |
Affymetrix Human Transcriptome Array 2.0 ST arrays were hybridized according to Affymetrix recommendations using the Affymetrix WT Plus kit. Affymetrix GeneChip® Human Transcriptome 2.0 ST microarrays (HTA2) were hybridized with 5.5 ug of labeled DNA.
|
| Scan protocol |
HTA 2.0 ST arrays were scanned using the Affymetrix 3000 7G scanner and the Command Console software version 4.1.2.
|
| Data processing |
CEL files were first processed using the Affymetrix® Expression Console Software (build 1.3.1.187) to produce CHP files for each of the eight samples. The Gene Level - Default: RMA-Sketch normalization was used. The eight CHP files were then imported into the Affymetrix® Transcriptome Analysis Console (TAC) 2.0 (build 2.0.0.9) using the Gene Level Differential Analysis option. Within TAC, the genome assembly hg19 was used with the annotation file HTA-2_0.na34.hg19.transcript.csv. Intensity values reported for each transcript cluster ID are calculated as log2 of Tukey's bi-weight average.
|
| |
|
| Submission date |
Aug 26, 2014 |
| Last update date |
May 06, 2015 |
| Contact name |
Eric Christian Rouchka |
| E-mail(s) |
eric.rouchka@louisville.edu
|
| Organization name |
University of Louisville
|
| Department |
Biochemistry and Molecular Genetics
|
| Lab |
KY INBRE Bioinformatics Core
|
| Street address |
522 East Gray Street
|
| City |
Louisville |
| State/province |
Kentucky |
| ZIP/Postal code |
40292 |
| Country |
USA |
| |
|
| Platform ID |
GPL17586 |
| Series (1) |
| GSE60760 |
Gene expression responses to chronic low dose arsenite exposure |
|
