小鼠4-细胞胚胎OPS法冷冻保存技术  

Vitrification of Mouse 4-Cell Embryos by OPS Method

作　　者：周光斌[1] 金方[1] 侯云鹏[1] 杨中强[1] 刘国世[1] 田见晖[1] 朱士恩[1] 

机构地区：[1]中国农业大学动物科技学院,北京100094

出　　处：《中国兽医学报》2006年第2期210-212,共3页Chinese Journal of Veterinary Science

基　　金：北京市重点科技项目(H022020060420);北京市自然科学基金资助项目(6012011)

摘　　要：在25℃室温和37℃恒温台条件下，利用玻璃化冷冻溶液EFS30、EFS40、EDFS30或EDFS40，对小鼠4-细胞胚胎进行玻璃化冷冻保存．以解冻后培养72h的囊胚发育率为其体外发育能力的考核指标，同时对解冻后培养1～3h的胚胎进行移植以判定其体内发育潜力。开放式拉长塑料细管（OPS）二步法冷冻保存，即胚胎首先移入预处理液（10％EG或10％EG＋10％DMSO）中平衡30s，再移入玻璃化溶液中洗涤后吸入OPS管中，分别经35、30或25s后直接投入液氮中冷冻保存。一步法冷冻保存则无需预处理液处理。结果表明，小鼠胚胎4-细胞一步法和二步法冷冻后囊胚最高发育率分别为87．7％和88.6％，与对照组（93.0％）差异不显著（P〉0．05）。利用最佳冷冻组获得的143枚胚胎移植于12只假妊娠50～60h的受体母鼠，结果有4只妊娠产仔17只，妊娠产仔率为42．5％（17／40），与对照组59．4％（19／32）差并不显著（P〉0.05）。The experiments were conducted at 37℃ to cryopreserve mouse 4-cell embryos by vitrification with EFS30, EFS40, EDFS30 and EDFS40 （an ethylene glycol-based vitrification solution） respectively in open pulled straw （OPS）. In the 1-step OPS method, embryos were vitrified after direct exposure to vitrification solutions for 15s, 25s or 35s, while in the 2-step method, the superovulated embryos were first pretreated to either 10 % Ethylene glycol （EG） or 10 % EG ＋ 10 % DMSO for 30s respectively, then exposed to a vitrification solution （VS） for 25s, 30s or 35s and then immersed in liquid nitrogen. After warming, the survival of embryos was assessed by their ability to develop to blastocysts or develop to term after transfer. The higher blastocyst rates of vitrified embryos were from EDFS30 group in the 2-step OPS method （88. 6% ） and EDFS40 group in the 1-step method（87.7%）, which were similar （P〉0. 05） to that （93.0%） obtained in non-vitrified fresh embryos. The embryos derived from the best vitrified group or fresh 4-cell embryos （used as control） after culture for 1 to 3 h were transferred to 50-60 h pseudo-pregnant female mice. 17 fetuses were obtained from 4 of 12 recipients which had received 143 vitrifiedwarmed embryos. In the pregnant recipients, the percentage of transferred embryos developed to young in the treated group （42. 5 % ） and control （59.4 % ） showed no difference （P〉0. 05）. These results demonstrated that mouse 4-cell embryos could survive cryopreservation by vitrification in OPS.

关 键 词：OPS法 玻璃化冷冻 4-细胞胚胎 小鼠 

分 类 号：S814.8[农业科学—畜牧学]