Abstract: In recent years, with the development of the grouper industry, hybrid breeding has played an important role in the development and cultivation of new species of grouper farming. The karyotype analysis of fish is of great significance in fish hybrid breeding, genetic relationship and genetic variation. Fisheries Research Institute of Fujian carried out research on the hybrid of Epinephelus akaara ♀ and E. moara ♂ in 2013, and successfully obtained the hybrid F1 (E. akaara ♀ × E. moara ♂), but no chromosome karyotype analysis of hybrid F1 has been reported. Thus, using E. akaara as female parent and E. moara as male parent, we obtained hybrid generation of normal development by artificial insemination technology, which called “Red cloud grouper”. In this study, the karyotype of F1 groupers (E. akaara ♀ × E. moara ♂) was studied. The juvenile fish were intraperitoneally injected with bovine serum albumin (BSA), followed by phytohemagglutinin (PHA) and colchicine solutions. The metaphase chromosome spreads were obtained from head kidney cells. The results of the study showed that the number of chromosomes of the hybrid F1 groupers (E. akaara ♀×E. moara ♂) was 48 (i.e. 2n=48), of which one pair was metacentric chromosomes (m), one pair was sub-metacentric chromosomes (sm), and two pairs were sub-telocentric chromosomes (st) and twenty pairs were telocentric chromosomes (t). Among these 48 chromosomes of the hybrid F1 groupers (E. akaara ♀×E. moara ♂), the largest relative length was (5.15±0.28), and the smallest relative length was (2.84±0.11). The number of chromosome arms (NF) of hybrid F1 groupers (E. akaara ♀×E. moara ♂) was 56. According to the analysis, karyotype formula of hybrid F1 groupers (E. akaara ♀×E. moara ♂) was 2n=48=2m+2sm+4st+ 40t, NF=56. So far, karyotypes of 29 species of groupers have been reported. The chromosome number of these groupers is 2n=48, and the chromosome number of the hybrid F1 groupers (E. akaara ♀×E. moara ♂) in this paper was consistent with this. It can be seen that 2n=48 may be the most basic karyotype feature of groupers. During the evolution of groupers, the number of chromosomes is highly conserved. However, the karyotypes of different species are significantly different. The same grouper species may also have different karyotypes obtained by different researchers due to different geographic regions, different chromosome preparation methods or operating errors. In this study, the number of chromosomes of the hybrid F1 of E. akaara ♀ and E. moara ♂ was the same as that of the two parents, but the chromosome karyotypes were quite different from those of the two parents. The chromosomes karyotypes could not all be found in the two parents. Maybe, a complex chromosomal mutation had occurred. This research may provide an important scientific basis for the genetic relationship analysis, genetic variation of the hybrid F1 groupers (E. akaara ♀ × E. moara ♂), as well as the germplasm identification, character selection and germplasm improvement of the hybrid offspring.