Due to their special phylogenetic position in the Euarchontoglires and close affinity to primates, tree shrews have been proposed as an alternative experimental animal to primates in biomedical research. However, the population genetic structure of tree shrews has largely remained unknown and this has hindered the development of tree shrew breeding and selection. Here we sampled 80 Chinese tree shrews (Tupaia belangeri chinensis) in Kunming, China, and analyzed partial mtDNA control region sequence variation. Based on our samples and two published sequences from northern tree shrews (T. belangeri), we identified 29 substitutions in the mtDNA control region fragment (~604 bp) across 82 individuals and defined 13 haplotypes. Seventeen samples were selected for sequencing of the cytochrome b (Cyt b; 1134 bp) gene based on control region sequence variation and were analyzed in combination with 34 published sequences to solidify the phylogenetic pattern obtained from control region data. Overall, tree shrews from Kunming have high genetic diversity and present a remarkable long genetic distance to the two reported northern tree shrews outside China. Our results provide some caution when using tree shrews to establish animal models because of this apparent genetic difference. In addition, the high genetic diversity of Chinese tree shrews inhabiting Kunming suggests that systematic genetic investigations should be conducted before establishing an inbred strain for medical and biological research.