De novo assemblies of transcriptome sequence of F(1) interspecies hybrids between Mandarin fish Siniperca chuatsi (Basilewsky) (♀) and Golden mandarin fish Siniperca scherzeri Steindachner (♂) were generated using Illumina sequencing. In the present study, 64 microsatellite primer pairs were designed on basis of the transcriptome data. Those primer pairs were then characterized both in S. scherzeri and S. chuatsi. Thirty-eight polymorphic loci were detected in 37 individuals from a wild S. scherzeri population with 2-10 alleles per locus. The values for observed and expected heterozygosities ranged from 0.0000 to 0.8378 and from 0.4235 to 0.8771, respectively. Meanwhile, 36 microsatellite loci were polymorphic in 30 individuals from a wild S. chuatsi population with 2-8 alleles per locus. High cross-species transferability was detected. These markers will facilitate further studies on the conservation genetics, population structure and construction of high-density linkage map. What's more, they will be useful for parental selections in controlled hybridization breeding programs.