The nerippe fritillary butterfly, Argynnis nerippe , is listed as an endangered species in Korea. Establishment of effective conservation strategies can be aided by the development and application of molecular markers that can be used to investigate the population genetics of the butterfly. Therefore, in this study, we identified ten microsatellite markers specific to A. nerippe using the Next-Seq 500 platform, and applied these markers to investigate the characteristics of five South Korean butterfly populations. Genotyping of 48 A. nerippe individuals from five localities showed that at each locus the number of alleles ranged from 4 to 14, and that the observed and expected heterozygosities were 0.324–0.863 and 0.138–0.985, respectively. Significant deviation from the Hardy–Weinberg equilibrium was not observed at any locus. Population structure analysis indicated that there are two genetic groups in Korea, but no population-based gene pool assignments were found. Analysis of FST, RST, and a principal coordinates analysis suggested that the Gureopdo and Yaecheon populations were isolated from other populations. Genetic isolation of the Gureopdo population may be a consequence of unequal population change between Gureopdo and inland populations and to the offshore habitat of Gureopdo. Genetic isolation of the Yaecheon population may be a consequence either of the southernmost location of the population or of the limited sample size available. Further studies with increased sample sizes will be necessary to draw robust conclusions on population isolation and to devise conservation strategies.