Genespring gx software version 14

RNA was extracted from PBMCs using the RNeasyMini Kit (Qiagen, Hilden, Germany). Fifty ng of total RNA extracted were amplified and fluorescently labelled RNA amplification was performed using Low Input Quick Amp Labelling Agilent Kit (Agilent, Santa Clara, CA, USA) according to the manufacturers' instructions. Labelled RNA were purified using RNeasyMini Kit (Qiagen, Hilden, Germany), samples were vacuum dried at 55 °C and resuspended in the hybridization mixture containing hybridization buffer and blocking reagent. Hybridization was carried out on the S. scrofa V2 4x44k microarray (Agilent) in Agilent hybridization oven at 65 °C for 17 h according to the manufacturer's instructions. Following the hybridization, microarrays were washed with GE Wash Buffer 1 and GE Wash Buffer 2 and scanned with Agilent G2505 microarray scanner (Agilent Technologies, Santa Clara, CA, USA). The results were extracted using Feature Extraction software version 11.0 and analyzed using GeneSpring GX software version 14.5 (Agilent Technologies, Santa Clara, CA, USA). Data were normalized using quantile shift and reduced with fold-change (FC) cut-off set to 1.25 and then analyzed with Friedman and a posteriori Wilcoxon tests.