Abstract
This study was performed as part of the effort to develop artificial seed production techniques for stone flounder Kareius bicoloratus, which has a great potential for farming in northeast Asian countries. To develop a sperm cryopreservation protocol that can be applied for artificial fertilization, we carried out experiments on the key factors for successful fish sperm cryopreservation, including cryoprotective agents (CPAs), diluents, equilibration times, and dilution ratios. The CPAs tested in this study were dimethyl sulfoxide (DMSO), glycerol, methanol, and ethylene glycol, and the diluents were glucose and sucrose. The equilibration times tested in the experiments ranged from 0 to 210 s and the dilution ratios ranged from 1:1 to 1:1000. According to the results of our experiments, the optimal CPA for stone flounder sperm cryopreservation was DMSO in concentrations ranging from 7.5% to 10% when 300 mM glucose was used as diluent and DMSO in concentrations ranging from 15% to 17.5% when 300 mM sucrose was used as diluent. Post-thaw sperm motility was high, with equilibration times up to 150 s, and dilution ratios up to 1:10 (semen:CPA + diluent) showed no significant differences in the effect on post-thaw sperm motility. The cryopreserved sperm showed more significant DNA damage than the fresh sperm with the severity of DNA damage being in inverse proportions to post-thaw sperm motility and survival rate. It is expected that these results will be useful for applications of artificial fertilization at stone flounder hatchery farms.
Published Version
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