Área de Genética y Reproducción Animal, Centro de Biotecnología Animal, SERIDA, Gijón (Asturias), Spain.
Theriogenology. 2012 Oct 15;78(7):1465-75. doi: 10.1016/j.theriogenology.2012.06.018. Epub 2012 Aug 24.
In this work we analyzed the effects of three culture systems on developmental ability of bovine embryos in vitro produced with sexed sperm, the survival to vitrification (cryologic vitrification method) of such blastocysts, and their pregnancy rates after embryo transfer to recipients, both as fresh and after vitrification/warming. Finally, we measured the accuracy of the sorting protocol by a polymerase chain reaction-based method to validate the embryo sex at blastocyst stages. We confirmed an individual effect of the bull as well as development rates of embryos produced with sorted sperm lower than embryos with unsorted sperm, independent of the culture system used. The cryoresistance to vitrification of embryos produced with sexed sperm did not differ from that of conventionally produced embryos (re-expansion rates at 24 and 48 h: 74.6% vs. 75.5%, and 64.5% vs. 68.1% for embryos produced with conventional and sorted sperm, respectively; hatching rates at 48 h: 63.55% vs. 55.5% for embryos produced with conventional and sorted sperm, respectively). Finally, no significant differences were found in pregnancy rates after the embryo transfer of fresh and vitrified/warmed blastocysts (52.8% vs. 42.0%, respectively; P > 0.05). Male and female embryos produced with sorted sperm showed the same quality in terms of developmental ability, cryoresistance, and pregnancy rates after transfer. Our culture system, coupled with the vitrification in fiber plugs, provides good quality sex-known embryos which survive vitrification at similar rates than embryos produced with conventional unsorted sperm; also it produces good pregnancy rates after transfer of sexed embryos both fresh and after vitrification and warming.
在这项工作中，我们分析了三种培养系统对用性别鉴定精子体外生产的牛胚胎发育能力的影响，对这种囊胚进行玻璃化冷冻保存（玻璃化冷冻方法）的存活率，以及将胚胎新鲜或玻璃化/解冻后移植到受体后的妊娠率。最后，我们使用聚合酶链反应（PCR）方法对分选方案的准确性进行了测量，以验证囊胚阶段胚胎的性别。我们证实了公牛的个体效应以及用分选精子生产的胚胎的发育率低于未分选精子生产的胚胎，而与使用的培养系统无关。用性别鉴定精子生产的胚胎对玻璃化冷冻的抗逆性与常规生产的胚胎没有差异（24 和 48 小时的再扩张率：用常规和分选精子生产的胚胎分别为 74.6%和 75.5%，64.5%和 68.1%；48 小时的孵化率：用常规和分选精子生产的胚胎分别为 63.55%和 55.5%）。最后，新鲜和玻璃化/解冻胚胎移植后的妊娠率没有显著差异（分别为 52.8%和 42.0%；P>0.05）。用分选精子生产的雄性和雌性胚胎在发育能力、抗逆性和移植后妊娠率方面表现出相同的质量。我们的培养系统与纤维塞中的玻璃化相结合，提供了高质量的性别已知胚胎，其玻璃化冷冻保存存活率与用常规未分选精子生产的胚胎相似；并且，在新鲜或玻璃化/解冻后移植性别鉴定胚胎后，也能获得良好的妊娠率。
