【作者】 贾世全；

【导师】 王杏龙；

【作者基本信息】 扬州大学 ， 动物遗传育种与繁殖， 2007， 硕士

【摘要】 本文主要就不同分割液、培养液、分割处理方法及不同的冷冻保护液和的冷冻处理方法对小鼠桑椹胚和囊胚的影响进行了系统的研究。结果如下:1首先研究了不同的分割液和不同胚胎发育阶段对分割效果的影响,发现在12.5%蔗糖分割液中分割桑椹胚时,分割成功率显著高于m-PBS处理组(60.83% vs 51.67%,P<0.05 ),而分割囊胚时两分割液之间无显著性差异(P >0.05 );在m-PBS中分割囊胚和桑椹胚,其成功率前者显著高于后者(70.37%vs 51.67%,P<0.01),但是在12.5%蔗糖分割液中分别对囊胚和桑椹胚进行分割时,两者无显著性差异(P>0.05)。2无论是桑椹胚还是囊胚,其半胚经不同培养液m-PBS和TCM-199-Hepes培养后存活率差异均不显著(62.50% vs 63.64%;80.36% vs 82.76%,P>0.05);桑椹胚和囊胚分割后的半胚经m-PBS培养液培养后,所得半胚存活率两者间差异显著(62.50% vs 80.36%,P<0.05),经TCM-199-Hepes培养液培养后,所得半胚存活率两者间差异显著(63.64% vs 82.76%,P<0.05)。3桑椹胚去致密化后,其分割成功率显著高于未处理组(73.08% vs 51.92%,P<0.01),半胚培养后的囊胚发育率也显著高于未处理组(71.15% vs 46.15%,P<0.01)。4半胚经琼脂包埋后,利用VSⅠ和VSⅡ冷冻液进行OPS法玻璃化冷冻,解冻后其存活率均显著高于未处理组(90.16% vs48.28%;88.52% vs 44.83%,P<0.01),培养48h后的半胚发育率,处理组也均显著高于未处理组(61.82% vs 25.86%;61.11% vs 24.13%,P<0.01)。琼脂包埋组解冻后的半胚存活数和发育率,VSⅠ和VSⅡ之间差异均不显著(90.16% vs 88.52%;61.82% vs 61.11%,P>0.05);未包埋组解冻后VSⅠ和VSⅡ之间差异也均不显著(48.28% vs 44.83%;25.86% vs 24.13%,P>0.05)。5桑椹胚和囊胚分别经两种不同保护液(VSⅠ和VSⅡ)冷冻后形态良好率差异均不显著(96.94% vs 95.96%;86.87% vs 84.38%,P>0.05);经VSⅠ保护液冷冻后,桑椹胚和囊胚之间形态良好率差异非常显著(96.94% vs 86.87%,P<0.01),经VSⅡ保护液冷冻后,两者之间形态良好率差异也极显著(95.96% vs 84.38%,P<0.01)。6小鼠胚胎经分割后冷冻,其平均存活率为40.48%,经冷冻后分割的平均存活率为81.63%,两者之间差异非常显著(P<0.01)。更多还原

【Abstract】 In this paper, the effect of different splitting-solutions, medium, splitting method, different refrigerating fluid and freezing method on the morulae and blastulae of mice were studied. The main results were as follows:1 First studied the effect of different splitting-solutions and different stage of embryo on the splitting. The success rate of morula splitted in 12.5% sucrose was higher than splitted in m-PBS(60.83% vs 51.67%,P<0.05), while there’s no significant difference when the blastulae were splitted(P>0.05). Splitted the blastulae and morulae in m-PBS, the success rate showed significant difference (70.37%vs 51.67%,P<0.01), but there’s no significant difference when they were splitted in 12.5% sucrose (P>0.05).2 No matter morulae or blastulae, the livabilily of half-embryos between the m-PBS and TCM-199-Hepes showed no significant difference (62.50% vs 63.64%;80.36% vs 82.76%,P>0.05); while cultured in m-PBS, there was significant difference between morulae and blastulae(62.50% vs 80.36%,P<0.05), and so as TCM-199-Hepes(63.64% vs 82.76%,P<0.05).3 The success rate of decompacted morulaes was higher than untackled(73.08% vs 51.92%,P<0.01), and there was significant difference in the development rate of demi-morulae (71.15% vs 46.15%,P<0.01)4 Demi-embryos were vitrified with VSⅠand VSⅡby the way of the OPS, after thawed, the livability of demi-embryos embed by agar was higher than untreated group in both two vitrified medium(90.16% vs48.28%;88.52% vs 44.83%, P<0.01), after cultured 48h, the developmental rate of demi-embryo about treated group was higher than that untreated group (61.82% vs 25.86%;61.11% vs 24.13%,P<0.01). In the group of agar , there was no significant difference between the VSⅠand VSⅡof livability and developmental rate(90.16% vs 88.52%;61.82% vs 61.11%,P>0.05), so as the group without tackled (48.28% vs 44.83%;25.86% vs 24.13%,P>0.05).5 No matter VSⅠor VSⅡ, after vitrification, there was no significant difference between the rate of morphologically of morulae and blastulae(96.94% vs 95.96%;86.87% vs 84.38%,P>0.05).Vitrifide with VSⅠ, there was significant difference between the morphologically rate of morulae and blastulae (96.94% vs 86.87%,P<0.01),and so as the VSⅡ(95.96% vs 84.38%,P<0.01).6 Vitrified the splitted embryos of mice, the average livability was 40.48%, and the average livability of splitted embryo after vitrification was 81.63%, there showed significant difference between them (P<0.01).更多还原