To investigate the applicability of microsatellite markers developed from Megalobrama amblycephala in M.pellegrini and find the molecular markers to identify the M.amblycephala,M.pellegrini and their reciprocal hybrids,90 pairs of M.amblycephala EST-SSR primers were synthesized for cross-species amplification on the genome DNA of M.pellegrini,and 26 pairs of microsatellite primers of M.amblycephala and M.pellegrini were screened for PCR amplification on the genome DNA of M.amblycephala,M.pellegrini and their reciprocal hybrids.The results revealed that 76 pairs(84.4%)of the primers were amplified successfully.Fifty pairs were screened from these 76 primers to test the polymorphism in M.pellegrini population and 13 markers were polymorphic with an average of 3.2 alleles per locus,the mean observed heterozygosity being 0.72 and the mean PIC being 0.42.The results of PCR amplification on the genome DNA of M.amblycephala,M.pellegrini and their reciprocal hybrids indicated that 4 primer pairs had no amplified products on M.amblycephala,and 2 primer pairs had no amplified products on M.pellegrini.Among the 20 loci which had effective amplified products,9 specific loci could identify M.amblycephala,M.pellegrini and the reciprocal hybrids,respectively.Combination of the other 17 specific loci also distinguished M.amblycephala,M.pellegrini and the reciprocal hybrids effectively.However,the reciprocal hybrids［M.amblycephala(♀)×M.pellegrini(♂),M.pellegrini(♀)×M.amblycephala(♂)］could not be identified through these markers used in this study.