|
| Status |
Public on Dec 01, 2008 |
| Title |
HFF 12hpi Rep 3 |
| Sample type |
RNA |
| |
|
| Source name |
human foreskin fibroblast (HFF) cell line; harvested after 12 hpi
|
| Organisms |
Homo sapiens; Human herpesvirus 5 strain Towne |
| Characteristics |
human foreskin fibroblast (HFF) cell line
|
| Treatment protocol |
Cells were synchronized in G0 by serum starvation for 60h (8), trypsinized and replated onto 100mm dishes at a density of 1x106 cells per plate for microarray and RT-PCR. Cells were then infected with the HCMV Towne strain 3 h after reseeding at an MOI of 1 for the microarray studies. Virus containing media was replaced by fresh media 3 h post infection (hpi), the cells were harvested by trypsinization after 12 or 24 hpi and the pellets were stored at -80°C until use.
|
| Growth protocol |
Primary human foreskin fibroblast cells were grown in Eagle’s minimal essential medium (MEM, Gibco BRL) supplemented with 10% fetal bovine serum (FBS, Atlanta Biologicals), 200U/ml penicillin, 200µg/ml streptomycin, 2mM glutamine and 1.5µg/ml Amphotericin B (all Gibco BRL). The telomerase immortalized human lung fibroblast (THF) cell line was grown in DMEM/F12 (Gibco BRL) supplemented with 10% FBS, 200U/ml penicillin, 200µg/ml streptomycin and 2mM glutamine. THF derived p53-/- telomerase immortalized human fibroblasts were grown in DMEM supplemented with 7.5% FBS, 200U/ml penicillin, 200µg/ml streptomycin and 2mM glutamine (6). All cells were maintained at 37°C in a humified atmosphere containing 5% CO2.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
SV Total RNA Isolation Kit (Promega).RNA was amplified with the Superscript RNA Amplification kit (Invitrogen)
|
| Label |
Cy5
|
| Label protocol |
Amplified RNA was labelled with Cy5 using the ULS labelling Kit for Combimatrix Arrays (Kreatech Biotechnology)
|
| |
|
| Hybridization protocol |
protocols for hybridization and stripping were provided by Combimatrix.
|
| Scan protocol |
The fluorescence of each feature on the hybridized slides was read on an Axon GenePix 4200A (Axon, Sunnyvale, CA), and the raw fluorescence values were obtained using the Combimatrix MicroArray Imager
|
| Description |
HFF 12hpi Rep 3
|
| Data processing |
First, a mean value was calculated for each feature on the array from the three technical replicates. Then, the variance stabilization normalization was calculated seperately for the 12 hpi and the 24 hpi values.
|
| |
|
| Submission date |
Nov 03, 2008 |
| Last update date |
Nov 03, 2008 |
| Contact name |
Holger Hannemann |
| E-mail(s) |
holgerh@uidaho.edu
|
| Phone |
2088859237
|
| Organization name |
University of Idaho
|
| Department |
MMBB
|
| Lab |
Lee Fortunato
|
| Street address |
PO Box 443052
|
| City |
Moscow |
| State/province |
ID |
| ZIP/Postal code |
83843 |
| Country |
USA |
| |
|
| Platform ID |
GPL7474 |
| Series (1) |
| GSE13437 |
p53 regulates viral gene expression during infection with human cytomegalovirus |
|
