Analyzing recessive mutations typically requires the production of three generations: an original heterozygous carrier, an F1 cross, and multiple F2 crosses. This can be shortened to two generations by producing uniparental progenies from the heterozygous carrier through gynogenesis. Gynogenesis is typically performed using UV-irradiated zebrafish sperm, but this limits its use to females carrying an easily identifiable recessive mutation, so as to recognize any contamination from sperm that was not deactivated. This could be overcome by using UV-irradiated heterologous sperm that is capable of initiating development but, if inactivation is not complete, results in either inviable or distinguishable hybrids. Koi carp Cyprinus carpio was evaluated as a candidate for this purpose.
Hybridization was performed by artificially crossing 18 zebrafish females with one koi carp male. Embryonic development was successfully initiated by koi carp sperm. There was high progeny mortality in all tested pairs over the first 24h and complete mortality prior to the swim-up stage. Several distinct and consistent deformities were observed in the hybrid embryos: blastomeres of unequal size, short bodies, and edema.
Gynogenesis was performed using UV-irradiated koi carp sperm and either an early pressure or a heat shock treatment was used to restore diploidy. Survival to 21 days post fertilization was similar between progenies from the two types of shocks, averaging 2.2% and 2.5% for early pressure (n=3) and heat shock (n=5), respectively. Thus far, only males have been present in gynogenetic progenies produced (n=8). Samples of males resulting from 2 separate early pressure treatments were spawned naturally with non-sibling females. Control males (half-siblings of the gynogenetic males) were also spawned with the same females. The proportion of males that were fertile, fertilization rates and survival from fertilization to swim-up larvae was recorded. Fertilization rates were significantly lower than the control in one gynogenetic progeny (p<0.10), but not in the second. Survival from fertilization to the swim-up stage was not significantly different from the control in either progeny. This demonstrates that gynogenetic males could be used as broodstock for the rapid development of homozygous mutant lines. Males from a third early pressure shock progeny and from five heat shock progenies will be tested for fertility and these results will be presented.