The HPV16-immortalized keratinocytes had been transfected with a plasmid carrying the genome of the human papillomavirus, HPV16 (Mitrani-rosenbaum et al., 1992, Intervirology33: 76-85). Early passage (passage 20, E) represents cells that underwent about 50 doublings after transfection.
Growth protocol
Cells were maintained in Keratinocyte Growth Medium (KGM): DMEM, 25% Nutrient mixture F-12 (HAM), 10% fetal bovine serum, 5 µg/ml insulin, 0.4 µg/ml hydrocortisone, 10−10 M cholera toxin, 10 ng/ml epidermal growth factor, 1.8×10−4 M adenine, 5 µg/ml transferrin, 2×10−9 M T3, 100,000 U/L penicillin, 100 µg/L streptomycin, 0.1 mg/ml amphotericin.
Extracted molecule
total RNA
Extraction protocol
Three independent cultures of Early cells were grown. RNA was isolated using RNeasy kits (Qiagen) according to the manufacturer's instructions.
Label
biotin
Label protocol
Standard Affymetrix probe synthesis for gene expression profile ( carried out in the microarray unit of the Weizmann institute)
Hybridization protocol
Standard Affymetrix hybridization protocol for gene expression profile ( carried out in the microarray unit of the Weizmann institute)
Scan protocol
Standard Affymetrix protocol for gene expression profile ( carried out in the microarray unit of the Weizmann institute)
Description
Human U133A 2.0
Data processing
The gene expression data were analyzed using the Microarray Suite (MAS) 5.0 algorithm. For each probe, a CHP output file contained a signal quantitative metric, which represents the relative level of expression of a transcript, and a “detection” classification of each signal as present, marginal, or absent.
