Restriction endonuclease cleavage patterns of mitochondrial DNAs (mtDNAs) from four species of jungle fowl were examined. The sequence divergence between the various mtDNA haplotypes was estimated from the numbers of common and different restriction fragments. The sequence divergence between Gallus gallus and respectively, G. sonnerati, G. varius, and G. lafayettei was 0.9%, 10.5%, and 12.6%. Assuming that the rate of evolution of mtDNA is 3% per million years (Myr), we estimated that the divergence of mtDNA of G. gallus from that of G. sonnerati, G. varius, and G. lafayettei occurred 0.3 Myr, 3.5 Myr, and 4.5 Myr ago, respectively. These maternal phylogenetic relationships are in good agreement with those inferred from allozyme data and reported in the literature.