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1.
试验研究了不同培养体系对猪体外成熟卵母细胞电激活后的体外发育的影响。试验1比较了猪孤雌发育胚胎在NCSU 23、G 2.2添加氨基酸(G 2.2-aa)和G 2.2培养体系中的体外发育效率,结果表明,3个组卵裂率差异不显著(P>0.05),NCSU 23体系的囊胚率显著高于其他2组(14.37±3.77 vs.2.67±2.68和3.13±0.45,P<0.01),但3组间囊胚细胞数差异不显著(21.29±5.27 vs.19.33±2.54和20.27±4.72,P>0.05)。试验2探讨了胚胎培养72h后更换培养液对孤雌发育胚胎的影响,结果显示,培养液更换对卵裂率和囊胚细胞数的影响差异不显著(P>0.05),但更换组囊胚率显著下降(12.50±1.49 vs.5.56±1.89、6.25±1.13、4.64±1.56,P<0.05)。试验3研究了在G 2.2-aa添加胰岛素、亚牛磺酸和半胱氨酸对孤雌激活胚胎发育的影响,结果显示,以上添加剂对卵裂率和囊胚细胞数无显著影响(P>0.05);在G 2.2-aa添加胰岛素、亚牛磺酸和半胱氨酸能明显提高囊胚发育率(6.38±1.00、6.20±2.08、8.73±1.03 vs.2.99±2.21,P<0.05),但以上各组的囊胚率明显低于NCSU 23体系(P<0.05)。结论:NCSU 23是猪孤雌激活发育胚胎较为理想的培养液。 相似文献
2.
作为葡萄糖同分异构体的果糖同样可以作为胚胎早期发育的代谢底物,而且可能在桑椹胚发育阶段之前就为胚胎发育所利用,有利于早期胚胎的体外发育。本实验结合囊胚细胞双重染色方法研究了在SOFaaci培养基中,果糖代替葡萄糖对牛早期胚胎体外发育效果的影响。结果表明:对第8天囊胚双重染色时,以Triton X-100处理60~69 s为宜,内细胞团(TCM)细胞数和囊胚细胞总数分别为48.2±12.4和142.5±30.1,二者之比约为0.33;果糖组与葡萄糖组卵裂率(62.2±3.87 vs 60.7±2.89)、囊胚率(32.0±1.73 vs 29.7±4.16),TCM细胞数(43.6±14.5 vs 40.2±11.3)和滋养层(TE)细胞数(99.3±26.7 vs 89.9±24.8)差异也不显著(P>0.05)。但是数据显示,果糖组囊胚细胞总数要高于葡萄糖组(142.3±21 vs 136.2±35)(P>0.05),说明果糖可以代替葡萄糖作为早期胚胎体外培养的能量代谢底物,而且可以提高囊胚质量。 相似文献
3.
牛磺酸对牛体外受精早期胚胎发育的影响 总被引：1，自引：0，他引：1
研究了牛体外受精后早期胚胎体外发育时向其基础培养液中加入牛磺酸对胚胎桑椹胚率、囊胚率和孵化囊胚率的影响.试验1:以TCM-199 10?S为基础培养液(对照组),再加入7、14 mmol/L的牛磺酸(试验组),试验组与对照组的桑椹胚率分别为48.1%、47.4%和43.2%;囊胚率分别为26.4%、22.3%和21.0%;孵化囊胚率分别为21.8%、18.7%和0.试验2:IVF后2细胞、4～8细胞及8～16细胞期,在基础培养液中分别添加7 mmol/L的牛磺酸时,桑椹胚率分别为48.7%、57.1%和52.0%,囊胚率分别为25.1%、30.7%和27.9%,孵化囊胚率分别为25.9%、28.6%和25.0%;而添加14 mmol/L牛磺酸时,桑椹胚率分别为50.4%、56.4%和55.4%,囊胚率分别为26.5%、31.3%和27.7%,孵化囊胚率分别为27.5%、31.1%和29.9%.结果表明,体外发育培养液中添加7 mmol/L牛磺酸可显著提高桑椹胚率和囊胚率(P＜0.05),并且在4～8细胞期添加14 mmol/L牛磺酸最为合适. 相似文献
4.
5.
利用放射自显影技术检测了小鼠着床前胚胎(4-细胞、8-细胞、桑葚胚和囊胚)不同发育阶段(早、中、晚)S期和M期卵裂球的比率，并研究了低温(4℃)对小鼠胚胎发育及DNA合成的影响。结果显示，早期和中期4-细胞胚胎S期卵裂球比率较高，分别是75%、92%，没有M期卵裂球，晚期4-细胞胚胎S期卵裂球比率较低(30%)，M期卵裂球比率为11%；早期和中期8-细胞胚胎S期卵裂球比率仍很高，分别是73%、80%，M期卵裂球分别为0、2.5%，晚期8-细胞胚胎S期卵裂球比率增至64%，M期卵裂球减少至9%；早、中、晚期桑葚胚的S期卵裂球比率分别是77%、72%和79%，M期卵裂球的比率分别是5%、3%和3%；早、中、晚期囊胚的S期卵裂球比率分别是78%、74%和77%，M期卵裂球的比率分别是4%、3%和4%。4℃的低温对小鼠早期胚胎的发育和DNA合成都没有明显影响；4-细胞、8-细胞胚胎和桑葚胚经低温处理及培养后其囊胚形成率和平均卵裂球数分别是62%和(26±6)个、54%和(25±7)个、87%和(34±8)个；4-细胞、8-细胞胚胎、桑葚胚和囊胚经低温处理后S期卵裂球比率分别是97%、77%、70%和74%。 相似文献
6.
采用端粒酶重复序列扩增(telomeric repeat amplification protocol,TRAP)-银染法,对小鼠卵母细胞、体内受精胚胎及重构胚的各阶段端粒酶活性进行测定,对PCR产物的电泳条带进行灰度值分析,计算端粒酶总产品总量。结果表明:(1)体内受精胚胎及核移植重构胚各阶段的端粒酶活性随胚胎分裂呈上升趋势,囊胚期最高(P0.05);(2)体内受精胚胎的端粒酶活性高于体细胞核移植重构胚端粒酶活性。同时进行囊胚计数并测定单卵裂球相对端粒酶活性,结果表明:单卵裂球的端粒酶活性呈下降趋势,囊胚期端粒酶活性最低(P0.05)。以上结果说明小鼠卵母细胞及体细胞核移植重构胚端粒酶活性变化与细胞发育水平及细胞发育全能性有关。 相似文献
7.
牦牛卵母细胞的体外成熟、种间受精与胚胎培养 总被引：1，自引：0，他引：1
探讨了卵母细胞体外成熟时间、卵母细胞质量、精子准备和受精卵培养体系对牦牛卵母细胞种间体外受精效果的影响。结果表明:牦牛卵母细胞随体外成熟培养时间延长,第一极体排出率增加,囊胚发育率以成熟培养24 h最高;A级卵母细胞种间受精后囊胚的发育率(48.77±3.76)%显著高于B级(32.05±5.24)%和C级(7.54±7.18)%(P<0.05);BO液洗涤离心处理的精子受精后卵裂率(82.53±6.54)%显著高于Percoll液分离精子受精后的卵裂率(67.39±4.50)%(P<0.05),而两者囊胚率差异不显著((42.32±4.13)%vs(35.59±5.62)%,P>0.05);荷斯坦奶牛精子浓度在1×106～5×106/mL范围与牦牛卵母细胞受精效果差异不显著。卵丘细胞、输卵管上皮细胞共培养和SOF液培养种间受精卵,其卵裂率差异不显著,但共培养组的桑葚胚、囊胚和孵化胚发育率均显著高于SOF液培养组。 相似文献
8.
通过聚合嵌合法初步建立一套黄牛和水牛种间嵌合的程序与方法。聚合嵌合法采用链酶蛋白酶消化透明带或用机械剥离法去除透明带，然后在含有100 μg/ml PHA的培养液中聚合形成嵌合胚。结果发现， 8-细胞黄牛胚胎聚合水牛桑椹胚与黄牛囊胚聚合水牛桑椹胚相比，聚合胚存活率和囊胚发育率均无显著差异（P>0.05）。采用显微手术法分离黄牛和水牛8-细胞胚胎卵裂球进行聚合，聚合率为92.3%，囊胚发育率为58.3%，与用0.25%链酶蛋白酶分离胚胎卵裂球进行胚胎聚合的聚合率（86.7%）和囊胚发育率（46.2%）均无显著差异（P>0.05）。以上结果表明：①水牛和黄牛胚胎通过卵裂球聚合获得的种间嵌合胚胎能继续发育；②胚胎聚合前的发育阶段对其聚合成功率和随后的胚胎发育无明显影响。③胚胎卵裂球的分离方法（显微手术法和酶消化法）对其聚合率和囊胚发育率无明显影响。 相似文献
9.
中药成分小檗碱对小鼠2-细胞胚胎体外培养及移植效果的研究 总被引：3，自引：0，他引：3
以添加抗菌素的mCZB液为对照组,筛选比较了中药有效成分小檗碱不同浓度对小鼠2-细胞胚胎体外培养效果的影响,并计数所培养的孵化胚胎细胞数目,观察中药有效成分对胚胎细胞数目增殖的作用;通过将体外培养发育的囊胚进行移植,进一步验证其对小鼠胚胎移植效果和产仔发育的影响。结果表明:小檗碱最佳浓度为0.10μg/mL,120h孵化胚胎发育率和孵化胚胎细胞数目(89.9%,83.7±9.10)极显著高于对照组(50.7%,69.5±7.14)(P<0.01);小檗碱组培养囊胚移植妊娠率(66.7%)和离窝成活率(55.8%)均显著高于对照组(50.0%,45.2%)(P<0.05),其组间产仔率、初产仔鼠平均体重、离窝小鼠平均体重无显著差异(P>0.05)。其结果说明,小檗碱对体外胚胎生长发育和细胞增殖有促进作用,对胚胎附植及初生仔鼠无不良影响。 相似文献
10.
本试验对影响绵羊体外受精的因素进行了分析。结果表明:在相同条件下添加和不添加VEGF(血管内皮生长因子)的卵裂率分别为81.5%、84.9%(P>0.05),但囊胚发育率分别为31.5%和22.4%,差异显著(P<0.05)。受精时间8 h和12 h对卵裂率(80.8%、83.9%;P>0.05)和囊胚发育率(28.8%、23.5%;P>0.05)没有显著影响。卵巢采回后立即切割和在25~30℃的生理盐水中保存3 h的卵裂率分别为82.9%、86.5%,囊胚发育率分别为30.4%、29.7%,差异均不显著(P>0.05)。 相似文献
11.
Ulloa Ulloa CM Yoshizawa M Yamashita A Hama S Mitsui A Hashi C Abe H Hoshi H Fukui E Matsumoto H 《The Journal of reproduction and development》2008,54(6):465-472
The present study was conducted to determine the criteria for selecting good quality embryos on Day-2 post-insemination and at the blastocyst stage. Bovine oocytes were matured, fertilized and cultured in vitro. First, Day-2 embryos were classified based on the number of blastomeres into 2-cell, 3- to 4-cell, 5- to 8-cell and >8-cell stage embryos; chromosome samples were then prepared. In the second experiment, the Day-2 embryos classified according to the number of blastomeres were cultured separately for an additional 6 days (Day 8). The resultant Day-8 blastocysts from each group of Day-2 embryos were classified into the following 3 grades based on morphology and diameter: Grade A, hatched and hatching blastocysts; Grade B, expanded blastocysts; and Grade C, unexpanded blastocysts. Chromosome samples were then prepared. The 5- to 8-cell stage Day-2 embryos had the lowest incidence of chromosomal abnormalities (13.5%, P<0.05) and the highest development rate to blastocysts (59.2%, P<0.05). Furthermore, the blastocysts derived from the 5- to 8-cell stage embryos had the largest mean number of cells (102.8+/-42.4, P<0.05), largest number of metaphases per blastocyst (9.5+/-4.8, P<0.05) and lowest incidence of chromosomal abnormalities (24.6%, P<0.05). The Grade A blastocysts had the largest mean number of cells (136.6+/-33.4, P<0.05), a large number of metaphases per blastocyst (11.9+/-5.5, P<0.05) and a low incidence of severe chromosomal abnormalities (17.3%). The results showed that, at Day 2, the 5- to 8-cell stage embryos were of better quality since they had the lowest incidence of chromosomal abnormalities and the highest blastocyst rate and the resultant blastocysts had the largest number of cells and lowest incidence of chromosomal abnormalities. In particular, selection of Grade A blastocysts can improve the development rate to term. 相似文献
12.
昆明小鼠早期胚胎体外发育阻滞原因分析 总被引：3，自引：0，他引：3
为了分析小鼠早期胚胎在体外发育阻滞的原因 ,建立早期胚胎体外培养系统 ,应用几种不同的培养系统对昆明小鼠单细胞胚胎在体外培养了 96～ 12 0 h。结果表明 ,小鼠单细胞胚胎在 M1 6 和 BWW培养液中卵裂均受到阻滞 ,且两者之间差异不显著 (P>0 .0 5 ) ;CZB和 HTF培养液能有效地克服小鼠胚胎的 2 -细胞发育阻滞 ,与 M1 6 和 BWW相比 ,2 -细胞卵裂率和囊胚率均存在显著性差异 (P<0 .0 1) ;在 M1 6 培养液中添加牛磺酸对早期胚胎发育有促进作用 (P<0 .0 1) ;小鼠早期胚胎在 CZB和牛输卵管上皮细胞 (COEC)共培养体系中的卵裂率较对照组明显提高 (P<0 .0 5 ) 相似文献
13.
Yan CL Yang QE Zhou GB Hou YP Zhao XM Fan ZQ Liu MQ Liu L Zhu SE 《Acta veterinaria Hungarica》2008,56(2):245-253
The present study was designed to investigate the cryotolerance of in vitro fertilised (IVF) mouse embryos at various preimplantation developmental stages. IVF mouse embryos were vitrified by the open-pulled straw (OPS) method. After warming, embryos were morphologically evaluated and assessed by their development to blastocysts, hatched blastocysts or term. The results showed that a high proportion (93.3-100.0%) of vitrified embryos at all developmental stages were morphologically normal after recovery. The developmental rate of vitrified 1-cell embryos to blastocyst (40.0%) or hatched blastocyst (32.7%) or term (9.3%) was significantly lower than that from other stages (P < 0.05). Vitrified embryos from 2-cell to early blastocyst stage showed similar blastocyst (71.8-89.5%) and hatched blastocyst rates (61.1-69.6%) and could develop to term without a significant loss of survival compared with those of fresh embryos (P > 0.05). Vitrified 2-cell embryos showed the highest survival rate in vivo (50.6%, 88/174), compared with that from other stages (9.3-30.5%, P < 0.05). The data demonstrate that the OPS method is suitable for the cryopreservation of IVF mouse embryos from 2-cell stage to early blastocyst stage without a significant loss of survival. Embryos at the 2-cell stage had the best tolerance for cryopreservation in the present study. 相似文献
14.
Glucose is utilized for oxidation and synthesis of various lipids in cultured rat embryos. The present experiment examined the effect of insulin on the incorporation of glucose into lipid fractions in rat embryos in vitro . Embryos at the 2-cell, 8-cell and blastocyst stages were incubated for 5 h in hamster embryo culture medium (HECM)-1 containing 14 C-glucose and 170 nmol/L insulin, or in HECM-1 containing only 14 C-glucose, and the oxidation of glucose in these embryos was examined. In addition, the total lipids of blastocysts were separated by thin layer chromatography and the radioactivity of the separated lipid fractions was measured. Oxidation of glucose was significantly increased after insulin treatment compared with that without insulin treatment in 8-cell embryos and blastocysts ( P < 0.05), but not in 2-cell embryos. Incorporation of glucose into lipids in blastocysts was significantly lowered by insulin treatment compared with that without insulin treatment ( P < 0.05). Most of the radioactivity was recovered from triacylglycerols of blastocysts and the remaining radioactivity was found in other neutral lipids and phospholipids. We conclude that insulin accelerates the utilization for oxidation of glucose and inhibits the storage of triacylglycerols in rat blastocysts. 相似文献
15.
This study was conducted to improve the developmental ability of nuclear transfer (NT) embryos by using blastomeres from in vitro fertilized (IVF) embryos with high quality as donor cells. The IVF embryos selected at the 2-cell stage at 24-h postinsemination (hpi) and again at the ≥8-cell stage at 48 hpi (Selected-IVF-embryos) showed the highest blastocyst formation rate among embryos. When blastomeres from the Selected-IVF-embryos (Selected-NT group) or Nonselected-IVF-embryos (Non-selected-NT group) were used as donor cells for NT, the blastocyst formation rate in the Selected-NT group (25.6%) was significantly higher than that in the Non-selected-NT group (13.5%). When blastomeres from the Selected-IVF-embryos at 108 (contained many cells before cell division) and 126 hpi (contained many cells immediately after cell division) were used as donor cells for NT (108- and 126-NT groups, respectively), the 126-NT group showed a significantly higher blastocyst formation rate (32.1%) than the 108-NT group (16.8%). Embryo transfer of blastocysts in the 126-NT group showed that 11 of 23 recipients became pregnant; nine calves were obtained. For the NT embryos reconstructed using in vivo derived embryos, 9 of 20 recipients became pregnant; seven calves were obtained. These results indicate that the blastocyst formation rate of NT embryos can be improved by using blastomeres from IVF embryos selected at the early developmental stage, especially immediately after cell division, and that the resultant NT embryos have a high developmental ability to progress to term that is comparable to NT embryos reconstructed using in vivo derived embryos. 相似文献
16.
Ulloa Ullo CM Yoshizawa M Komoriya E Mitsui A Nagai T Kikuchi K 《The Journal of reproduction and development》2008,54(1):22-29
The present study was conducted to determine the relationship between embryonic development speed at different stages (the cleaved stage at 52 h and the blastocyst stage at 6 days post insemination) and incidences of chromosome abnormalities in in vitro produced porcine embryos. Porcine oocytes were collected from 3-6-mm ovarian follicles obtained at a slaughterhouse and matured in modified NCSU-37 medium for 44-46 h. Following in vitro fertilization with a final concentration of 1 x 10(5) sperm/ml for 3 h, all oocytes were cultured in vitro for 52 h. Day-2 (52 h after insemination) embryos were classified according to their cleaved stages into 2-cell, 3- to 4-cell, 5- to 8-cell, and >8-cell stages; these were cultured separately for additional 4 days (Day 6). The resultant Day-6 blastocysts were classified according to the morphological diameter into 3 grades: Grade A, expanded blastocysts; Grade B, expanding blastocysts; and Grade C, early blastocysts. They were then analyzed chromosomally. The 3- to 4-cell and 5- to 8-cell embryos had significantly high blastocyst development rates (46.1 and 36.9%, respectively), and these blastocysts contained significantly more cells (40.2 and 42.4 cells, respectively) than those derived from 2-cell embryos and >8-cell embryos (28.6 and 26.5 cells, respectively). The incidence of chromosomal abnormalities was significantly higher in the blastocysts derived from 2-cell and >8-cell stage embryos than in the blastocysts derived from the other stage embryos. Furthermore, the grade A blastocysts had the lowest incidence of chromosomal abnormalities (35.3%) and contained the most cells (48.7 cells). Porcine in vitro production (IVP) yielded a high blastocyst rate and an excellent embryo quality when 3- to 4-cell and 5- to 8-cell stage embryos were selected on Day 2 after insemination. The same criteria yielded a higher quality of expanded blastocysts based on the stage of embryo development and morphology. 相似文献
17.
Thanh Quang Dang‐Nguyen Nguyen Viet‐Linh Tamas Somfai Seiki Haraguchi Bui Xuan Nguyen 《Animal Science Journal》2014,85(5):517-523
In the present study, we examined the development to blastocysts of large and small blastomeres from unevenly cleaved 2‐cell embryos (uneven 2‐cell embryos) in pigs. Proportion of blastocysts derived from large blastomeres (52.8 ± 6.4%) was significantly higher (P < 0.05) compared with small ones (32.1 ± 4.6%). However, there were no differences in total cell number, inner cell mass (ICM) cell number and ICM/total cells ratio between them. Of 53 sister blastomere pairs in the same embryos examined there were 12 pairs (22.6%) in which both blastomeres developed to blastocysts, 16 pairs (30.2%) in which only large blastomeres developed to blastocysts, and five pairs (9.4%) in which only small blastomeres developed to blastocysts. Relative total amount of active mitochondria in small blastomeres were lower (P < 0.05) than that of large blastomeres and blastomeres from evenly cleaved 2‐cell embryos. However, there was no difference in relative density of active mitochondria in these three types of blastomeres. In conclusion, blastocysts derived from small and large blastomeres in uneven 2‐cell embryos had comparable quality in terms of cell number, ICM number, ICM/total cell ratio and distribution of active mitochondria. The results suggest that these blastomeres may contribute multiple offspring production in pigs. 相似文献
18.
本研究通过线粒体分子探针标记技术检测孤雌激活早期胚胎线粒体的分布变化,运用实时荧光定量PCR技术检测mtDNA拷贝数的变化,揭示早期胚胎发育过程中线粒体分布、mtDNA拷贝数变化趋势。结果表明,成熟卵母细胞电激活后,由2-细胞胚胎开始,卵裂球内线粒体分布均匀且密集,每个细胞均有分布,卵裂球之外的空隙未见线粒体分布,直到囊胚形成,线粒体均有分布。孤雌激活4-细胞胚胎mtDNA拷贝数显著高于8-细胞胚胎mtDNA拷贝数(907210.77±145520.77,186224.33±103308.00,P<0.05),但显著低于2-细胞胚胎、桑椹胚、囊胚的mtDNA拷贝数(1563422.54±224666.51、1697626.25±176999.53和1752301.29±101146.64,P<0.05)。孤雌激活扩张囊胚mtDNA拷贝数最高,为2812545.67±156819.31,显著高于其他发育时期胚胎的mtDNA拷贝数(P<0.05)。由此可见,孤雌激活早期胚胎发育进程中线粒体分布及mtDNA拷贝数会发生变化。 相似文献
19.
The objective of this study was to compare the effect of two culture media: modified synthetic oviductal fluid (mSOF) and G1.2/G2.2, on the developmental competence of bovine somatic cell–cloned embryos. Cloned embryos were produced by transferring adult skin fibroblasts into enucleated MII oocytes. After activation, the reconstructed embryos were randomly allotted to either mSOF or G1.2/G2.2 for culture (the embryos were transferred from G1.2 to G2.2 on days 3 of culture). The development competence of cloned embryos in these two culture systems was compared in terms of cleavage rate, blastocyst formation rate and apoptosis cell number in day 7 blastocyts. To investigate the in vivo developmental competence of cloned embryos in the two culture systems, a total of 87 and 104 blastocysts derived from mSOF and G1.2/G2.2 medium groups were transferred individually to recipient Angus cows, respectively. No differences were observed in terms of cleavage rate, day 7 blastocyst rate and blastocyst cell number between these two culture systems. However, the day 6 blastocyst formation rate was significantly higher in G1.2/G2.2 than that in mSOF. In addition, blastocysts cultured in mSOF have a higher percentage of apoptotic blastomeres compared to those in G1.2/G2.2 (8.5 ± 1.2 vs 16.8 ± 1.5, p < 0.05). Although difference in pregnancy rate was not observed 40 days after embryo transfer, significantly higher pregnancy rate was observed in G1.2/G2.2 group after 90 days of embryo transfer (12.4% vs 37.5%, p < 0.05). Moreover, calving rate was significantly improved in G1.2/G2.2 group compared to mSOF group (27.9% vs 6.7%, p < 0.05). In conclusion, our results indicate that G1.2/G2.2 can improve developmental competence of bovine SCNT embryos both in vitro and in vivo, which is more suitable for culture of bovine SCNT embryos than mSOF medium. 相似文献