Genetic diversity of endangered Big-headed Turtle,28 individuals from Conghua,Guangdong and 21 individuals from Shouning,Fujian,was determined by using ISSR technique.There was high genetic diversity between the two populations and within the same population.The genetic diversity of Conghua population was higher than that of Shouning population.Genetic differentiation mainly occurred between populations (F_ST=0.371 5,G_st=0.205 9),but there was also gene flow between natural populations (N_m=1.927 9).UPGMA cluster analysis showed that the 49 individuals were divided into two clusters corresponding to the two populations.In addition,11 specific ISSR markers which can distinguish Conghua population from Shouning population were selected from 11 primers in two populations,using ISSR and Bulked Segregant Analysis (BSA).