We investigated the population structure of the spotted seal, Phoca largha, around Hokkaido, Japan, inferred from sequence variations of the mitochondrial cytochrome b gene. Twenty-six haplotypes were identified in 52 individuals, and 21 of them were newly found in the present study. To detect population structure, samples were divided into three groups—Mamiya, Okhotsk, and Habomai—based on breeding areas, and genetic difference among the groups was examined by exact test and estimation of Φ_ST and F_ST. The results of these analyses suggested that there were small but statistically significant genetic differences between the breeding areas.