Abstract: Using inter-simple sequence repeat (ISSR) analysis, the genetic diversity and the genetic structure of 7 natural populations of Trillium tschonoskii were investigated. Twelve ISSR primers generated highly reproducible and stable DNA fragments. Using these primers, 135 discernible DNA fragments were produced. Of these, 46 (PPB= 34.07%) were polymorphic loci, which indicated that low levels of genetic variation existed in the populations. The result of POPGENE analysis indicated that the level of genetic variation of T. tschonoskii (H_e=0.075 9,H_o=0.120 0) was lower than that of other endangered plants. The population in Wolong possessed the highest level of genetic diversity (PPB=18.52%,H_e=0.041 7,H_o=0.068 4), while that in Dabazi exhibited the lowest levels of genetic diversity (PPB=8.89%, H_e=0.022 0, H_o=0.034 8). A high level of genetic differentiation among populations which might be caused by barriers to gene flow and genetic drift was detected based on Nei's genetic diversity analysis(0.555 4) and the analysis of molecular variance (0.525 3). Restricted gene flow (Nm=0.400 2) among populations may result from several factors, such as low seed dispersal, geographical isolation of population and self_pollination. Based on ecological and genetic information available for T. tschonoskii, some conservation strategies are proposed.