|
| Status |
Public on Sep 20, 2009 |
| Title |
H5D, Rapamycin, Rep3 |
| Sample type |
RNA |
| |
|
| Source name |
H5D cells, 50nM Rapamycin
|
| Organism |
Rattus norvegicus |
| Characteristics |
rapamycin response: Intermediate
doubling time: 26.6h
tumorigenic: Yes
|
| Treatment protocol |
10^6 cells were plated on 100mm plates and allowed to attach overnight. Cells were treated with DMSO vehicle or 50nM rapamycin for 24 hours.
|
| Growth protocol |
GN5 cells were cultured in DMEM/F12 containing 10% fetal bovine serum, 2mM L-glutamine and 0.1% antibiotic-antimycotic solution. H5D cells were cultured in Waymouth MB 752/1 containing 10% fetal bovine serum and 0.1% antibiotic-antimycotic solution. All cells were grown at 37°C with 5.0% carbon dioxide.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Trizol extraction of total RNA was performed according to the manufacturer's instructions. Total RNA was further purified using the RNeasy kit (Qiagen). RNA quality was evaluated using the Agilent Bioanalyzer.
|
| Label |
Biotin
|
| Label protocol |
Biotinylated cRNA was prepared from 500ng of total RNA using the TotalPrep RNA Amplification kit (Applied Biosystems) according to the manufacturer's directions.
|
| |
|
| Hybridization protocol |
cRNA (750ng) was hybridized at 58°C for 16 hours on a RatRef-12 Expression BeadChips. BeadChips were washed and stained as outlined in the Illumina protocol.
|
| Scan protocol |
BeadChips were scanned using the Illumina Iscan.
|
| Description |
Derived from azo-dye induced hepatocellular carcinoma in Fischer rat
|
| Data processing |
Scanned files were quantile normalized using BeadStudio software (Illumina). The data was further analyzed using Partek software, version 6.3 (Partek). For gene expression comparisons, genes with detection p-values >0.01 from BeadStudio or fold-change less than 1.2 were excluded from further analyses. A two-sample t-test using p-value cutoff of 0.05 with multiple test correction (Bejamini and Hochberg false discovery rate) was applied for each gene to determine if the gene was differentially expressed in the pairwise comparison. Triplicate control and rapamycin samples from each cell line were used in these comparisons.
|
| |
|
| Submission date |
Aug 14, 2009 |
| Last update date |
Sep 14, 2009 |
| Contact name |
Jennifer Ann Sanders |
| E-mail(s) |
Jennifer_Sanders@brown.edu
|
| Organization name |
Rhode Island Hospital
|
| Department |
Pediatric Endocrinology
|
| Street address |
593 Eddy Street
|
| City |
Providence |
| State/province |
RI |
| ZIP/Postal code |
02903 |
| Country |
USA |
| |
|
| Platform ID |
GPL6101 |
| Series (2) |
| GSE17662 |
Modulation of gene expression by rapamycin in hepatic cell lines, H5D and GN5 |
| GSE17677 |
Modulation of gene expression by rapamycin in hepatic cell lines |
|
