subject: individual 4 melanoma: none cdkn2a: wild type mc1r: wild type twins: 1 cell type: keratinocyte disease state: none
Extracted molecule
total RNA
Extraction protocol
Total RNA isolation from primary cultures on passage 3-4 was performed using the Trizol extraction method (Invitrogen Life Technologies, Carlsbad, CA) followed by purification in commercial columns (Qiagen, Valencia, CA). Total isolated RNA was further purified using an RNeasy kit (Qiagen, Valencia, CA). RNA concentration was determined using a NanoDrop Spectrophotometer (Thermo Scientific) and integrity of the RNA was verified by Bioanalyzer 2100 (Agilent, USA). The RNA integrity number was in all cases higher than 8.
Label
Cy3
Label protocol
Overall, 50 ng of RNA were labeled using Low input Quickamp Labeling kit (Agilent, US).
Hybridization protocol
The hybridization, washing and staining protocols used in the study were those recomended by Agilent (USA). The One-color Microarray-based gene expression Analyisis protocol version 5.7.March 2008 were used in the study.
Scan protocol
The arrays were scanned using the DNA Microarray Scanner G2565CA (Agilent, US). Finally, Feature Extraction Software (FES, Agilent, USA) was used both to perform the quality control process and to extract the information.
