The red-spotted apollo butterfly, Parnassius bremeri Bremer, 1864 (Lepidoptera: Papilionidae), is an endangered species in South Korea. Development and application of molecular markers to assess population genetics perspectives can be used as a basis to establish effective conservation strategies. In this study, we developed 12 microsatellite markers specific to P. bremeri using Illumina paired-end sequencing and applied the markers to South Korean populations to understand population characteristics. Genotyping of 40 P. bremeri individuals from three localities showed that at each locus, the observed number of alleles ranged from 6 to 22 and the observed and expected heterozygosities were 0.500-1.00 and 0.465 to 0.851, respectively. Significant deviation from the Hardy？Weinberg equilibrium was not observed in all loci studied. The population based F ST and RST collectively suggest that at least the Samcheok population in northernmost Gangwon Province has a significant divergence from the remaining two populations (P < 0.01), and this result is also reflected in the forewing length. Further studies with an increased sample size will be necessary to draw robust conclusions and devise conservation strategies.