Red spotted grouper, Epinephelus akaara is a popular aquaculture species in many Asian countries. This species is a protogynous hermaphrodite that first differentiates into female and changes to male later. Due to this reproductive characteristic, stable supply of male and female gametes is a key to the success of seed production in this species. Thus, understanding early gonadal differentiation is required to develop effective sex control techniques. Red spotted grouper were reared in indoor tanks and sampled every 5 days from 40 days post-hatch (DPH) to 130 DPH. Changes of gonadal tissues were examined and analyzed by means of histology. A pair of gonadal primordium has already existed underneath the kidney in the posterior part of the body cavity at 38 DPH when this study began. Gonadal primordia of 38, 40 DPH consisted of germ cells surrounded by a few somatic cells. The blood vessel was observed in the gonadal primordium at 45 DPH. The number of somatic cells and size of gonadal primordium increased age-dependently up to 60 DPH. Formation of ovarian cavity was obvious by two protuberant aggregations of somatic cells at 65 DPH. Completed ovarian cavity and oogonia were first observed in the gonad of one fish sample at 105 DPH. Based on these histological observations, it can be suggested that induction of primary male differentiation could be more successfully applied at around 60 DPH in this species.

The sex differentiation of fishes occurs under the control of genetic and various environmental factors. DM-domain containing genes are novel zinc finger transcription factors and play key roles in sex determination. In order to isolate the wrasse DMRT (wDMRT) cDNA from the protogynous wrasse (Halichoeres tenuispinnis), the wrasse testis cDNA library was screened using the P-labeled PCR products, which were amplified with the degenerate primers from conserved DM-domain regions of several DMRT genes. Among a few positives obtained through screening, the full length wDMRT cDNA of 2.9kb size encoding a predicted 300 amino acid residues was isolated. The sequence analysis exhibited 60%, 43% sequence identity with rainbow trout and tilapia DMRT1, respectively. RT-PCR assay showed that wDMRT was expressed specifically in male testis. Also, wDMRT gene was strongly expressed in May during reproductive season, when the reproductivity of wrasse is most active. This results suggested that wDMRT gene function in testis differentiation The conserved DM-domain regions were amplified using PCR from DMRT genes of several species among Labridae, and their sequences were determined. The sequence of DM-domain region of Halichoeres. tenuispinis was identical to those of Pseudolabrus japonicus, Pteragogus flagellifera, and showed 94% identity with that of Halichoeres poecioptrerus.