Zhizhong Xiao

The fish species Oplegnathus fasciatus exhibits an X₁X₁X₂X₂/X₁X₂Y sex determination mechanism. This species holds considerable economic value and displays pronounced sexual dimorphism in growth. Therefore, the development of a rapid and accurate method for sex identification is critical to enhancing breeding efficiency and maximizing production value. Using third-generation PacBio whole-genome sequencing, we identified a homologous region in the samd3/elf3 intergenic region of the X and Y chromosomes of O. fasciatus. Analysis of the whole-genome sequence revealed a large DNA insertion marker fragment within this region. Using specifically designed primers, two bands of 390 bp and 1008 bp were successfully amplified in males, whereas only a single 390 bp band was detected in females. This marker can be easily distinguished by agarose gel electrophoresis, greatly enhancing the efficiency and accuracy of sex identification. This study not only expands the molecular marker system for sex identification of O. fasciatus but also offers a valuable methodological reference for sex identification in other economically important fish species. These findings have significant implications for germplasm improvement and efficient selection in aquaculture.

Spotted knifejaw (Oplegnathus punctatus) is an economically important marine cultured species exhibiting a unique complex sex chromosome system (X₁X₁X₂X₂ in females and X₁X₂Y in males), with males possessing one fewer chromosome (2n = 47) than females (2n = 48) and an abnormally large Y chromosome. Additionally, males demonstrate significant growth advantages over females. Rapid and accurate sex identification is essential for effective culture management, selective breeding, and population control. In this study, we identified a homologous region of the tmem88 gene containing large DNA insertion markers on the X and Y chromosomes through whole-genome sequencing of O. punctatus. The X₁ chromosome harbors a 278 bp DNA fragment, whereas the Y chromosome contains a 1472 bp fragment, resulting in a 1194 bp size difference indicative of structural variation in the non-coding region of the tmem88 gene. We developed a rapid detection method based on this variation, utilizing a pair of primers that amplify two distinct bands (278 bp and 1472 bp) in male (X₁X₂Y) individuals and a single 278 bp band in female (X₁X₁X₂X₂) individuals when analyzed by agarose gel electrophoresis. This method enables efficient and accurate sex differentiation in O. punctatus, significantly reducing the time required for identification and enhancing detection efficiency. This study provides a valuable tool for the rapid identification of sex in O. punctatus, facilitating improved breeding strategies and supporting the large-scale production of high-quality fry.