We detected and isolated thirty-two microsatellite loci from transcriptome-derived microsatellite DNA for Quasipaa boulengeri. The isolated loci were all polymorphic and the number of alleles per locus ranged from 2 to 10. The observed and expected heterozygosity per locus varied from 0.1493 to 0.9375 and from 0.1563 to 0.9688, respectively. By multiple comparisons with Bonferroni correction, seven loci were deviated from Hardy-Weinberg equilibrium and significant linkage disequilibrium was detected among twenty-five pairs of loci. Those variable microsatellite sites will be applied to assess gene flow between populations and investigate the evolutionary process of chromosomal polymorphism. Moreover, microsatellite loci were served as a valuable resource for drawing up criterion for the conservation management of this threatened species.