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ISSN : 1225-3480
본 연구에서는 굴, Crassostrea gigas의 계획적인 인공종묘생산 및 우량형질의 정자를 보존하기 위하여 냉동보존 시 적정결빙억제제 (cryprotective agent, CPA) 및 농도를 알아보고자, CPA 종류 및 농도별 독성 및 냉동보존 효과를 파악하고해동 후 정자의 세포 손상을 조사하였다. CPA 종류 및 농도별침지시간에 따른 굴 정자의 독성평가를 실시한 결과, 생존율및 운동성은 DMSO가 가장 좋았으며, 다음으로 EG,glycerol, Methanol 순이었다. 희석액으로 여과해수를 사용하여 CPA 종류 및 농도별 냉동보존 결과, 15% DMSO로 냉동보존 한 정자의 생존율 및 운동성이 가장 높았다. 굴의 냉동/해동 정자를 주사전자현미경으로 관찰한 결과, DMSO, EG,methanol, glycerol 순으로 세포 손상이 적었으며, 농도는15, 20, 10, 5% 순이었다. 이상의 결과를 종합한 결과, 굴 정자의 냉동보존 시 여과해수를 희석액으로 사용할 경우 적정CPA는 DMSO이며, 농도는 15%였다.
This study aims to find out a suitable cryoprotective agent (CPA) for cryopreservation and its optimum concentration in order to conduct planned artificial seed production of Pacific oyster, Crassostrea gigas and to preserve superior sperm. For this, we tried to understand toxicity and the effect of cryopreservation by CPA type and concentrations first and then looked into cell damage of the sperm after thawing. Toxicity analysis on the sperm of Pacific oyster according to different CPA and immersion time shows that dimethyl sulfoxide (DMSO) comes first when it comes to survival rate and mobility followed by ethylene glycol (EG), glycerol and methanol. To identify the optimum CPA and its level, filtered seawater was used as a diluent before cryopreservation for 30 days. As a result, cryopreserved sperm of Pacific oyster with 15% of DMSO showed the highest survival rate and activation. Also, we observed the cryopreserved and thawed sperm with Scanning electron micrographs by CPAs and concentrations. Consequently, DSMO showed the lowest cell damage followed by EG, methanol, glycerol and the level was 15, 20, 10, 5% respectively. In a nutshell, it is proven that the optimum CPA and its level is 15% of DMSO.
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