Genetic disturbances in wild populations of Medaka (Oryzias latipes) have resulted from the introduction of populations originating from the commercial orange-red strain (himedaka). The “himedaka” phenotype, caused by a defect in melanin deposition in skin cells, is conferred by a mutated recessive allele (b allele) on the slc45a2 locus (linkage group 12). To examine genetic introgression from “himedaka” to wild Medaka populations, a new DNA marker (b-marker) was constructed that detects fish with the b allele according to a length polymorphism of the promoter region between the wildtype (B) and b alleles. Among 169 fish from 45 wild populations in the Yamato River, Nara Prefecture, all specimens with the orange-red phenotype (from 7 populations) had a homozygous b/b genotype. Although most of the wildtype fish had a homozygous B/B genotype, 12 wildtype fish from 9 populations (3 of which consisted of fish with the orange-red phenotype) had the heterozygous genotype (B/b). These results indicated a high level of cryptic genetic introgression from “himedaka” to wild Medaka populations.