Striped beakfish Oplegnathus fasciatus and spotted knifejaw O. punctatus belong to the family Oplegnathidae, and are economically important fisheries resources in Japan and Korea. These species are distributed in the Pacific Ocean around southern parts of the Korean Peninsula, Japan, Taiwan, and Hawaii. These fish inhabit coastal rocky reefs, and feed on benthic animals. Mature females of these species spawn several times on the sandy/gravel bottom from April to July [1–3]. The ability to produce a hybrid, or to maintain reproductive isolation, is limited by two sets of reproductive barriers [4]. Prezygotic barriers include ecological isolation (e.g., geographical separation), temporal isolation (e.g., different mating seasons), behavioral isolation (e.g., incompatible courtship), mechanical isolation (e.g., incompatible genitalis), and gametic isolation (e.g., incompatible gametes). Postzygotic barriers include hybrid inviability, hybrid sterility, and hybrid breakdown [5]. Kamei and Takama [6] reported that O. fasciatus and O. punctatus would employ similar reproductive strategies (spawning ecology, such as spawning season, spawning time, spawning area, and spawning behavior), thus supporting the potential for hybridization. In the wild, fish with a similar body color pattern to artificially inseminated hybrids between O. fasciatus and O. punctatus [7, 8] were rarely found. Even though these two species have been speciated, why does the hybrid appear in the wild? To investigate fundamental information on the reproductive system of the hybrid between these two species, we explored possible prezygotic barriers to hybridization by comparing spontaneous spawning with artificial insemination. O. fasciatus, O. punctatus, and their F1 hybrids (O. fasciatus 9 O. punctatus) were reared in sea net cages (age 4–6 years) at Fisheries Laboratory, Kinki University, Wakayama, Japan. On 2–14 June 2007, we performed observation of spontaneous spawning in 3,000-L tanks in each cross group. The following four crosses were conducted: