|
| Status |
Public on Apr 06, 2020 |
| Title |
HPV16-transfected human foreskin keratinocytes_line A_timepoint 4_DAC treated |
| Sample type |
RNA |
| |
|
| Source name |
primary human foreskin keratinocytes transfected with HPV16
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: primary human foreskin keratinocytes transfected with HPV16
transformation stage: immortal
timepoint: timepoint 4
treatment: 5000 nM 5-aza-2’-deoxycytidine (DAC)
|
| Treatment protocol |
Cells were treated with either 5000nM 5-aza-2’-deoxycytidine (DAC; Sigma-Aldrich) dissolved in PBS or PBS only for 5 days. Medium with or without DAC was refreshed every day.
|
| Growth protocol |
Establishment and culture of the HPV16 (FK16A and FK16B) and HPV18 (FK18A and FK18B) transformed keratinocyte cell lines was described previously (Steenbergen et al., Oncogene 1996).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using TRIzol Reagent according to the manufacturer’s instructions (ThermoFisher Scientific).
|
| Label |
Cy3
|
| Label protocol |
500 ng of total RNA was reverse transcripted to cDNA using T7 Promoter Primer and MMLV-RT. Then the cDNA was converted to aRNA using T7 RNA polymerase, which simultaneously amplifies target material and incorporates cyanine 3- or cyanine 5-labeled CTP (Two-Color Microarray-Based Gene Expression Analysis Protocol, Agilent).
|
| |
|
| Hybridization protocol |
825ng of Cy3 and Cy5 labelled aRNA were hybridized competitively for 17h in a 65°C hybridization oven (G2545A, Agilent) set to 10rpm in a final concentration of 1X GEx Hybridization Buffer HI-RPM, according to the manufacturer's recommended protocol (Two-Color Microarray-Based Gene Expression Analysis, Agilent).
|
| Scan protocol |
Arrays were scanned at 5um resolution on an Agilent DNA Microarray Scanner (G2505B, Agilent) using the default settings for 4x44k format two-color arrays.
|
| Description |
raw data file: US22502676_251485074150_S01_GE2_107_Sep09_1_3.txt
none
FK16AT4+
|
| Data processing |
Images were auto gridded, analyzed and data extracted using Agilent Feature Extraction software. Raw expression data generated by the Feature Extraction software was imported into R using the LIMMA package in Bioconductor. The intensity distributions within and between arrays were normalized using the quantile scaling algorithm in LIMMA.
|
| |
|
| Submission date |
Sep 26, 2019 |
| Last update date |
Apr 06, 2020 |
| Contact name |
Renske DM Steenbergen |
| E-mail(s) |
r.steenbergen@amsterdamumc.nl
|
| Organization name |
Amsterdam UMC, location VUmc
|
| Department |
Pathology
|
| Lab |
Molecular Pathology
|
| Street address |
De Boelelaan 1117
|
| City |
Amsterdam |
| ZIP/Postal code |
1081 HV |
| Country |
Netherlands |
| |
|
| Platform ID |
GPL4133 |
| Series (2) |
| GSE138079 |
Identification of deregulated pathways, key regulators, and novel miRNA-mRNA interactions in HPV-mediated transformation. [mRNA cell lines-Agilent] |
| GSE138081 |
Identification of deregulated pathways, key regulators, and novel miRNA-mRNA interactions in HPV-mediated transformation. |
|
