New conditions for generation of gynogenetic half-tetrad embryos in the zebrafish (Danio rerio)

The generation of gynogenetic half-tetrads is an important tool for genetic mapping and mutant screens in zebrafish (Danio rerio). Half-tetrad gynogenesis can be accomplished using hydraulic pressure to disrupt microtubule-mediated segregation during meiosis II, which normally occurs after fertilization. However, the equipment used in the original studies is no longer available, and the reported pressures have not proven effective. Here, microsatellite markers were employed in a redetermination of the pressures required to generate zebrafish half-tetrads with currently available equipment. Two criteria were used to choose conditions for half tetrad gynogenesis: expected ploidy as confirmed with microsatellite markers and maximal egg viability. Pressures between 6140 and 6840 psi generate half- tetrad embryos, with optimal viabilities attained at 6140 and 6240 psi. As predicted, early pressure treatment of eggs fertilized with nonirradiated sperm created triploid embryos.