On the Miura Peninsula of central Honshu, Japan, there are international ports that, it is concerned, provide opportunities for the introduction of house mice via overseas cargoes. To evaluate the occurrence of such an introduction of overseas mice, in this study, we analyzed mitochondrial cytochrome b (Cytb) gene and the morphological characteristics of house mouse (Mus musculus) samples (n = 47) from five localities on the Miura Peninsula. All of the Cytb sequences of the present mouse samples were recognized as the musculus type corresponding to the subspecies musculus in Eurasia, which have been seen frequently in the Japanese Islands in previous studies. In addition, consistent with typical Japanese house mice, external morphology of the present mouse samples showed a shorter tail length and head and body length as compared with those of M. domesticus domesticus in Europe, M. musculus musculus in northern Europe and M. castaneus castaneus in southeastern Asia in Marshal (1998). Accordingly, in the current research area on the Miura Peninsula, it seems that there has been no influence by overseas mice, and the present mice examined are recognized as the typical Japanese house mice, considering the above Cytb properties and morphological characteristics.