We evaluated the genetic variation and genetic structure of northern largemouth bass(N), Florida largemouth bass, (F), and their obverse (N♀×F♂) and inverse (F♀×N♂) crosses, using 18 pairs of microsatellite primers. We generated between 2–8 alleles at each locus, with a mean allele number of 5.0. Six subspecies specific loci (Jzl48, ) could be used to distinguish parents and their reciprocal hybrids. The number of effect alleles (), and polymorphic information content were all highest in F♀×N♂ (3.199 70.570 6, respectively). The observed heterozygosity ( analysis of all groups and loci revealed that both reciprocal hybrids had lower genetic differentiation with N species. The UPGMA clustering analysis suggested that the offspring was nearest to the female brood stock for the obverse cross and nearest to the male brood stock for the inverse cross, suggesting that northern largemouth bass pass more genetic material than Florida largemouth bass.