To provide information for the management of sika deer in Tsurugi Quasi-National Park, Shikoku, Japan, we analyzed population genetic structure and migration patterns in the national park and surrounding areas based on from six to seven microsatellite loci using tissue and fecal samples. Bayesian clustering divided the deer into two populations, the Tokushima population and the Kochi population. Twenty-four of 28 samples on Mt. Tsurugi were assigned the Kochi population and the spatial boundary between the two populations was found in the foothills of the Tsurugi mountain range. These results suggest that the invasion of sika deer into Tsurugi Quasi-National Park may have started from the foothills of the Tsurugi mountain range by expansion of the Kochi population. Therefore, proper management of Kochi population is crucial in preventing further migration of sika deer into Tsurugi Quasi-National Park.