Evidence has shown that bacterial genomes have undergone random shuffling of genomic elements consisting of one to two genes. In order to delineate such genome-shuffling events in mammals, we constructed a high-resolution map of Sus scrofa chromosome 3 (SSC3) with a total of 116 genes/markers. Alignment of this pig map to orthologous regions in human, dog, mouse and rat led to the identification of 31 provisional conserved ancestral blocks (CABs) in these five species. Among them, only 3 CABs (<10%) had one gene, indicating that one-gene shuffling is not frequent in mammals. The sizes of CABs vary significantly within a species, but each may be relatively consistent in different species with a scale to species-genome evolution. The type and frequency of rearrangement events that takes place, either intra- or interchromosomal, depends on the evolutionary regions and species under comparison. Characterization of 36 tentative breakpoint regions flanking these 31 CABs indicated that they occupied approximately 43 Mb in length and featured genome deserts, gene duplications, and birth/death of species-specific genes in humans. Identification of CABs provides an alternative for further determination of the evolutionary make-up of mammalian genomes.