Characterisation and cross-amplification of 42 microsatellite markers in two Amphiprion species (Pomacentridae) and a natural hybrid anemonefish to inform genetic structure within a hybrid zone.

The hybrid anemonefish, Amphiprion leucokranos, is known to be the product of ongoing, introgressive hybridization between parent taxa Amphiprion sandaracinos and Amphiprion chrysopterus. Hybridization is an important evolutionary phenomenon contributing to biodiversity within marine systems, where hybrid zones provide ideal systems in which to study hybridization events. Here, a suite of 42 Amphiprion microsatellite markers (including development of 8 novel markers) were cross-amplified in individuals from parent taxa and hybrid populations to facilitate investigation into the relatedness of hybridizing species across the A. leucokranos hybrid zone. Analysis revealed 15, 20 and 24 highly polymorphic loci (PIC > 0.5) in the two parent species and hybrid, respectively, for use in population genetic and parentage studies, with 305 unique alleles found overall (ranging from 1 to 13 alleles per locus) and 7 alleles per locus on average. Observed and expected heterozygosities ranged from 0.000 to 1.000 and 0.000 to 0.978, respectively. Significant deviations from Hardy-Weinberg equilibrium were found in eight loci, possibly due to relatedness among samples or the presence of null alleles. Use of the suite of markers tested here will provide valuable insights into the contemporary population structure and introgression among species and hybrids within the Amphiprion leucokranos hybrid zone, as well as inform future ecological and evolutionary studies of anemonefishes more broadly.