Development of a multiplex microsatellite assay for parentage assignment in orange‐spotted groupers (Epinephelus coioides)

Establishment of reliable pedigree information of different lineages is crucial for selective breeding and stock enhancement programs. However, owing to the absence of a reliable assay for parentage assignment, such information is lacking in orange‐spotted grouper (Epinephelus coioides), which is an economically important coral‐reef fish. In this study, we developed two multiplex microsatellite polymerase chain reaction systems that consisted of 12 microsatellite markers (seven in system A and five in system B) for parentage assignment of orange‐spotted groupers. The theoretical exclusion probability per locus ranged from 0.079 to 0.436 when both parents were unknown (E‐1P) and from 0.191 to 0.614 when one parent was known (E‐2P). The combined exclusion probability values of 12 loci were 98.8% (E‐1P) and 100% (E‐2P), respectively. The accuracy of the parentage test for a mixed family (15 dams and 12 sires, 226 offspring) was 100%. In addition, two wild populations (79 individuals) were added as interferential offspring, only four of which were misassigned to the stock. The success rate of assignment was 98.69%. We provide a reliable microsatellite‐based assay for parentage assignment in orange‐spotted groupers, which will be useful for selective breeding and stock enhancement programs for this species and maybe other groupers.