摘要
为了研究适合于牛体外受精胚胎的一步除防冻剂的冷冻方法，特进行两个试验。试验１分别用１．５Ｍ甘油＋０．２５Ｍ蔗糖、１．５Ｍ乙二醇和１．５Ｍ丙二醇作防冻剂冷冻体外受精后第７天，已发育至囊胚阶段的牛胚胎。使胚胎降温至－７℃后，植冰，然后以０．３℃／分的速率降温至－３０℃，立即将胚胎投入液氮中冷冻保存。在３７℃水中解冻胚胎后，使其直接在含１５％胎犊血清（ＦＣＳ）的磷酸缓冲液（ＰＢＳ）中脱去防冻剂。经体外培养７２ｈ后，３组胚胎的孵化率分别为７７．２７％（１０２／１３２）、７３．２４％（１０４／１４２）、４７．９０％（５７／１１９），第１、２组的孵化率极显著高于第３组（Ｐ＜０．０１），说明用１．５Ｍ丙二醇溶液冷冻的胚胎，在解冻后不宜直接在ＰＢＳ中脱防冻剂。试验２比较用不同浓度（１．０Ｍ０１．５Ｍ，２．０Ｍ）的乙二醇和不同投液氮温度（－２５℃，－３０℃，－３５℃）冷冻胚胎的效果。结果表明乙二醇浓度和投液氮温度对胚胎孵化率均无显著影响（Ｐ＞０．０５），各组胚胎的孵化率变动于７４．４４％－８５．４８％之间。
Two experiments were conducted to study one step dilution mothods for frozen-thawed bovine in vitro embryos. In Exp. 1, Day 7blastoczysts were equilibrated for 15 min in 1.5 M glycerol (G)+ 0.25Msucrose(S), 1.5M ethylene glycol(EG) or 1.5M propylene glycol (PG) in PBS supplented with 15% FCS. The embryos loaded in 0.25 ml French straws were put into a freezer at -7℃,seeded 5 min later and cooled at the rate of 0.3℃/min to -30℃, at which temperature they were plunged into liquid nitrogen. After being thawed in water at 37℃ , the embryos were expelled directly into PBS/ FCS. They were cultured in vitro for 72 h. The hatching rates of the embryos frozen in 1.5M G+0. 25M S and 1.5M EG were 77.27%(102/132),73.24% (104/142) respectively, which were signifi- cantly higher than that (47.90%,57/119) of embryos frozen in 1.5M PG(P<0.01) In Exp. 2, the effect of EG concentration (1.0M, 1.5M and 2.0M) and plunge temperature (-25℃,-30℃ and -35℃) on embryo survival was examined in a 2x3 factorial design. Embryos were exposed to 1.0M, 1.5M or 2.0m EG in PBS/FCS for 15 min.They were cooled and seeded as in Exp. 1, and then cooled at the rate of 0.3℃/min to -25℃, -30℃ or -35℃ before being plunged into liquid nitrogen. Results after in vitro culture of the frozen-thawed embryos indicate that neither EG concentration nor plunge temperature affected the hatching rate of the embryos, which ranged from 74.44% 50 85.48%.
出处
《中国畜牧杂志》
CAS
北大核心
1996年第2期9-11,共3页
Chinese Journal of Animal Science
