Restoration of the loach, Misgurnus anguillicaudatus, from cryopreserved diploid sperm and induced androgenesis

In the present study, we evaluated the feasibility of using cryopreserved diploid sperm as a repository genebank for the loach, Misgurnus anguillicaudatus, along with a rederivation strategy utilizing induced-androgenesis. Firstly, we evaluated three types of media for egg inactivation: Hank's saline solution + 0.5% bovine serum albumin (BSA), Ringer's solution + 0.5% BSA, and masu salmon seminal plasma. Haploid and diploid sperm were taken from diploid and tetraploid loaches, respectively. Fresh and cryopreserved haploid or diploid sperm were then used to fertilize intact or UV-irradiated eggs from wild diploid females. The irradiation media evaluated here successfully maintained the egg quality over 2 h. Fertilization and hatching rates of eggs fertilized with cryopreserved diploid sperm were 11.68 ± 6.74% and 7.14 ± 6.29% respectively, compared to 63.51 ± 10.68% and 45.19 ± 16.2% for intact eggs fertilized by fresh haploid sperm. All-male inheritance was confirmed by determination of larval morphology, ploidy status and microsatellite genotypes of putative androgenetic progeny.