Tsoongiodendron odorum is an evergreen broad-leaved tree and at risk for becoming extinct in the wild. In this study, 13 T. odorum populations was analyzed using inter-simple sequence repeat (ISSR) markers to investigate the genetic diversity and structure. Eight primers produced discernible markers were applied. The percentage of polymorphic bands at the species and population level were 79.67% and 46.84%, respectively. Shannon’s phenotypic diversity index at the species and population level were 0.3880 and 0.2192, respectively. It indicates that T. odorum had a slightly higher level of genetic diversity than other related plants of the Magnoliaceae. The coefficient of genetic differentiation (ΦST) was 0.3752 by analysis of molecular variance (AMOVA) indicating that differentiation between populations was significant. Analysis of the ISSR data using STRUCTURE program and UPGMA revealed that the populations were clustered into three four population genetic clusters. The result of correlation analysis showed genetic distance was significantly positively correlated with geographical distance (DF=12，r=0.6635，p＜0.05). The Nankunshan, Bijiasha and Nongxiangshan populations should be given priority for conservation, because these populations had the highest genetic diversity among all the 13 populations investigated.