Gynogenesis and sex determination in Atlantic Halibut ( Hippoglossus hippoglossus)

Abstract

Gynogenesis refers to a process of uniparental inheritance whereby the resulting offspring retain only maternal DNA. It has been used to identify sex-determining mechanisms in fish and to produce all-female populations for aquaculture. The objective of this study was to develop a protocol for the production of gynogenetic Atlantic halibut for both these purposes. Various milt concentrations (1:20, 1:40 and 1:80 dilutions in seminal plasma) and UV doses (0–1382 mJ/cm 2) were tested for providing genetically inactivated, yet motile, spermatozoa for the production of gynogenetic haploids. Spermatozoon motility, assessed both as duration of swimming and percentage of motile spermatozoa, declined with increasing UV dose. Haploidy in developing embryos was determined either visually (‘haploid syndrome’) or by genetic analysis using microsatellite DNA. The optimum milt treatment for the production of haploids was 1:80 dilution followed by a UV exposure of 65 mJ/cm 2. Retention of the second polar body for the production of gynogenetic diploids was achieved by exposing newly activated eggs (15 min post-activation at 5–6 °C) to hydrostatic pressure of 8500 psi for 5 min. These treatments were combined to produce a population of gynogenetic diploids that was subsequently demonstrated to be comprised solely of females, thereby demonstrating that female is the homogametic sex in this species. It should therefore be possible to produce all-female populations of Atlantic halibut for commercial culture through gynogenesis or by crossing hormonally masculinized genotypic females to normal females.