Porcine Transcriptome Analysis by Microarray

Microarray analysis was performed as described previously [13 (link)]. Briefly, a GeneAtlas® system (Affymetrix, Santa Clara, CA, USA) was employed. RNA extraction and quality control were performed as described previously [13 (link)]. Labeled fragmented single-stranded cDNA (ss-cDNA) was synthesized by using purified total RNA (100-500 ng) as the template following Affymetrix WT PLUS Labeling Assay protocol. Then, the mixtures of biotinylate labeled ss-cDNAs were hybridized onto porcine Gene 1.1 ST Arrays (Affymetrix, Santa Clara, CA, USA). After being washed by a fluidic station, the arrays were scanned with an imaging station in a GeneAtlas® system (Affymetrix, Santa Clara, CA, USA), and the acquired array raw data were analyzed with the Affymetrix Command Console Software Version 1.4. Quantile normalization and subsequent data processing were performed by the Affymetrix Transcriptome Analysis Console (TAC) Software 4.0. The cutoffs for differentially expressed genes (DEGs) were set at fold change > 1.5 or <0.67 and FDR < 0.01. Gene Ontology (GO) (http://www.geneontology.org/) and Reactome pathway database (http://www.reactome.org) were used for further analysis.