The development of pig genome maps has rapidly progressed in recent years. This is largely due to the development o f genome mapping resources such as somatic cell hybrid panels, radiation hybrid panels and a number of reference populations. The objective o f this study was to further increase the marker density on pig genome maps using comparative mapping information and the novel technique of representational difference analysis (RDA). Information on conservation of synteny between the human and pig was used to generate markers surrounding the evolutionary breakpoint on human chromosome (HSA) 12, which is syntenic with pig chromosomes (SSC) 5 and 14. Seven gene markers were mapped using cytogenetic, radiation hybrid and genetic linkage mapping techniques. The recently described technique of RDA was also implemented to isolate genetic markers. RDA was performed using two pig populations divergent in genotype at the RYR1 locus and in phenotype for pork production traits. Eighteen markers were isolated using RDA, and most flank putative quantitative trait loci (QTL) for performance traits. Two of the isolated markers were homologous to the IRF6 and KCNB2 genes, which were subsequently characterized. The markers developed in this study further refine the human-pig comparative map and provide resources necessary for a better understanding of QTL for economically important pig production traits.