normalization data transformation protocol
The scanned images were processed using GCOS 1.4 (Affymetrix) according to the manufacturer. The resulting data sets (CEL files) were imported into ArrayAssist Expression Software (v5.2.0;Iobion Informatics LLC) and normalized using the PLIER algorithm in Array Assist to calculate relative signal values for each probe set. ID_REF = VALUE = Log2 PLIER signals
array scanning protocol
Signal intensities were detected by Affymetrix GeneChip Scanner 3000
The cRNA was hybridized to HG U133 plus 2.0 Arrays (Affymetrix) followed by washing and staining on the GeneChips Fluidics Station 450 (Affymetrix)
Double-stranded cDNA and biotin-labeled cRNA probes were made from 5 μg of total RNA by use of GeneChip® Expression 3' Amplification One-Cycle Target Labeling kit (Affymetrix).
nucleic acid extraction protocol
Trizol extraction followed by Rneasy clean up kit (qiagen) was performed according to the manufacturer´s instruction