Abstract: To guide breeding of Megalobrama terminalis, Megalobrama amblycephala and Erythroculter ilishaeformis, two hybrids Megalobrama amblycephala×Erythroculter ilishaeformis, Megalobrama terminalis×Erythroculter ilishaeformisand their parents were analyzed using sixteen microsatellite markers. The mean value of number of alleles (N_a) were 3.56, 3.63, 3.44, 4.00 and 4.31, respectively. The mean value of observed heterozygosity (H_o) were 0.3510, 0.3757, 0.3175, 0.3818, and 0.4079, respectively. The mean value of expected heterozygosity (H_e) were 0.6182, 0.6290, 0.5921, 0.6490 and 0.6825, respectively. And the average value of polymorphism information content (PIC) were 0.5354, 0.5367, 0.5258, 0.5785, and 0.6067, respectively. The average value of polymorphism information content (PIC) of two hybrid population were higher than their parent Megalobrama amblycephala, Megalobrama terminalis and Erythroculter ilishaeformis, indicating a high genetic diversity. The unweighted pair-group method with arithmetic means (UPGMA) showed that populations of the Megalobrama amblycephalaand Megalobrama terminalisfirst grouped together, and that Megalobrama amblycephala♀×Erythroculter ilishaeformis♂ andMegalobrama terminalis♀×Erythroculter ilishaeformis♂ first grouped together, and then they clustered with the Erythroculter ilishaeformis. In addition, the farthest genetic distance (0.5204) was between Megalobrama amblycephala and Erythroculter ilishaeformis, and the nearest genetic distance (0.0853) was between Megalobrama amblycephalaand Megalobrama terminalis. According to the genetic identity analysis, two hybirds groups showed the maternal effect. The alleles of two hybirds were from their parents based on genetic analysis. Particularly, microsatellite primers TTF3, TTF4, TTF10, and Mam25 identified 5 groups. Our results provide important value in breeding, germplasm resources preservation, and germplasm identification of Megalobrama terminalis×Erythroculter ilishaeformis andMegalobrama amblycephala×Erythroculter ilishaeformis.