2단계 급속동결 및 초자화 동결한 토끼상실배의 체외생존성에 관한 연구 = Studies on the Survival of Rabbit Morula Frozen by Two-Step Freezing and Vitrification

국문 초록 (Abstract)

본 시험은 수정란 급속동결보존기술의 개발을 위한 기초자료를 얻기 위하여 2단계 급속동결 및 초자화동결이 토끼 상실배의 체외발생등에 미치는 영향을 검토하고자 실시하였으며, 얻어진 결과는 다음과 같다.
1. 토끼 상실배를 1.5, 2.0, 2.5 및 3.0 M glycerol과 0.5 M sucrose가 포함된 동결액에 실온에서 10분간 노출후 -30℃에서 30∼40분간 정치하여 급속동결하였을 때 발생율은 각각 36.4, 83.3, 92.3 및 84.2%로 2.5 M glycerol에서 가장 높았다.
2. 토끼 상실배를 1.5, 2.0, 2.5 및 3.0 M 1,2-propanediol과 0.5 M sucrose가 포함된 동결액에 실온에서 10분간 노출 후 -30℃에서 30∼40분간 정치하여 급속동결하였을 때 발생율은 각각 26.6, 55.6, 65.0 및 52.9%로 2.5 M 1,2-propanediol에서 가장 높았으나, glycerol을 사용했을 때 보다 낮았다.
3. 토끼 상실배를 2.5, 3.0 및 3.5 M glycerol과 0.5 M trehalose가 포함된 동결액에 실온에서 10분간 노출후 액체질소에 침지하여 초급속동결한 결과 회수율은 각각 87.5, 92.5 및 92.5%, 형태적으로 정상인 수정란의 비율은 각각 37.5, 55.5 및 60.0%, 그리고 발생율은 각각 13.3, 36.4 및 37.5%로 3.5 M glycerol에서 가장 높았으나 초자화동결법보다 현저하게 낮았다.
4. 토끼 상실배를 25% glycerol과 25% 1,2-propanediol을 함유한 동결액에 실온에서 10분간 노출후 초자화동결했을 때 발생율은 75.0%로 실온에서 형평한 후 초자화동결이 가능하였다.

다국어 초록 (Multilingual Abstract)

This experiment was carried out to investigate on in vitro development of rabbit monla frozen by 2-step feezing and vitrification. The results obtained from this experiment are as follows;
1. When rabbit morula in m-PBS containing 1.5, 2.0, 2.5 or 3.0 M glyceral and 0.5 M sucrose for 10 min at room temperature were cooled at -30℃ for 30 to 40 min and plunged into liquid nitrogen, the proportion of embryo developed to expanded blastoyst was 36.4, 83.3, 92.3 and 84.2%, respectively. Glycerol 2.5 M showed higher survival than others.
2. When rabbit morula in m-PBS containing 1.5, 2.0, 2.5 or 3.0 M 1, 2-propanediol and 0.5 M sucrose for 10 min at room temperature were cooled at -30℃ for 30 to 40 min and plunged into liquid nitrogen, the proportion of embrye developed to expanded blastocyst was 26.6, 55.6, 65.0 and 52.9%, respectively. 1, 2-propanediol was less effective than glycerol.
3. When rabbit morula in m-PBS containing 2.5, 3.0 or 3.5 M glycerol and 0.5 M trehalose for 10 min at room temperature were plunged into liquid nitrogen and thawed rapidly, the recovery rate of embryo was 87.5, 92.5 and 92.5%, the proportion of morphologically normal embryo was 37.5, 55.5 and 60.0%, and the proportion of embryo developed to expanded blastocyst was 13.3. 36.4 and 37.5%, respectively. The proportion of embryo developed to expanded blastocyst was higher in vitrification than in plunging into liquid nitrogen.
4. When rabbit morula were frozen by vitrification in m-PBS containing 25% glycerol. 25% 1, 2-propanediol, the proportion of embryo developed to expanded blastocyst was 75.0%, the result suggested that rabbit embryos could be frozen by vitrification after equilibration at room temperature.