Tree shrews (Tupaia belangeri) are widely distributed in southeast Asia, mainly in Hainan Island and southwest regions of China. In the present study, PCR amplification of CO I gene sequence of 1398 BP was used to analyze 10 geographic populations consisting of 112 tree shrews as the research objects,. A total of 64 haplotypes were defined in CO I gene of the 112 samples, the haplotype diversity (Hd) averaged value was 0.9730, and the average value of nucleotide diversity (Pi) was 0.04494; analysis of AMOVA showed that variation among populations accounted for 93.09% of the total variance, suggesting that mutations occurred mainly in populations. The overall genetic differentiation index (F_ST) fixed to 0.93091, indicating that the populations of tree shrews have obvious genetic differentiation. Combination with neutral inspection and mismatch distribution analysis showed that T. belangeri had not experienced population expansion phenomenon in history . A phylogenetic tree constructed of haplotypes and NETWORK diagram showed that 10 the geographic populations of tree shrews were clustered into 4 branches: Hainan population, Daxin population, Pianma population and other populations. All of the results showed that there was higher genetic diversity of different geographical populations of the China in T. belangeri.  Different populations displayed obvious genetic differentiation and geographical barrier effects may be the main reason for the differentiation.