In this study, we determined how maternal dietary supplementation with pyridoxine combined with different sources of selenium (Se) affected global gene expression of porcine expanded blastocysts (PEB) during pregnancy. Eighteen gilts were randomly assigned to one of the three experimental diets (nZ6 per treatment): i) basal diet without supplemental Se or pyridoxine (CONT); ii) CONTC0.3 mg/kg of Na-selenite and 10 mg/kg of HCl-pyridoxine (MSeB 6 10); and iii) CONTC0.3 mg/kg of Se-enriched yeast and 10 mg/kg of HCl-pyridoxine (OSeB 6 10). All gilts were inseminated at their fifth post-pubertal estrus and killed 5 days later for embryo harvesting. A porcine embryospecific microarray was used to detect differentially gene expression between MSeB 6 10 vs CONT, OSeB 6 10 vs CONT, and OSeB 6 10 vs MSeB 6 10. CONT gilts had lower whole blood Se and erythrocyte pyridoxal-5-P concentrations than supplemented gilts (P!0.05). No treatment effect was observed on blood plasma Se-glutathione peroxidase activity (PZ0.57). There were 10, 247, and 96 differentially expressed genes for MSeB 6 10 vs CONT, OSeB 6 10 vs CONT, and OSeB 6 10 vs MSeB 6 10 respectively. No specific biological process was associated with MSeB 6 10 vs CONT. However, for OSeB 6 10 vs CONT, upregulated genes were related with global protein synthesis but not to selenoproteins. The stimulation of some genes related with monooxygenase and thioredoxin families was confirmed by quantitative real-time RT-PCR. In conclusion, OSeB 6 10 affects PEB metabolism more markedly than MSeB 6 10. Neither Se sources with pyridoxine influenced the Se-glutathione peroxidase metabolic pathway in the PEB, but OSeB 6 10 selectively stimulated genes involved with antioxidant defense.