形成两种胚细胞的

Our study includes four aspects. In the first aspect we study several important conditions of porcine oocytes maturation in vitro and oocytes cleavage after parthenogenetic activation and found mNCSU-23+15IU/mlPMSG+20IU/mlHCG+15% PFF+0.57mMcysteine is a good culture condition .When the Cocs are cultured in it ,the maturation rate and oocytes cleavage rate are higher than those of foreign covered. Our result are (86.7±3.35)% and (86.3±4.16)% and the highest report of foreign is(85.7±4.1)%.In the second aspect we study the effect of different chemical activations on development of porcine parthennogenetic embryo and found two best activation method. The first one is that putting the maturation MII oocytes in the 20μmol/L ionomycin for 30 minutes and then putting them in the NCSU-23 condition containing 5μg/mICB and 5mM/L6-DMAP for 3.5 hours, the oocytes cleavage rate and morulae/blastocysts development rate are (76.7±7.6)% and (37.1±6.4)%.The second one is that putting the maturation MII oocytes in the 200μM/L Thimerosal for 20 minutes and then putting them in the NCSU-23 condition containing 8mM DTT for 30 minutes

本研究分为4个部分，第一部分对影响猪卵母细胞体外成熟和孤雌激活后胚胎分裂的几个重要条件进行了比较研究，确立了一种较好的培养方法：与颗粒细胞共培养，找到了一种适合猪卵母细胞体外成熟的培养基：mNCSU-23+15IU/mlPMSG+20IU/mlHCG+15%PFF+0.57mM半胱氨酸，成熟率和分裂率分别为(86.7±3.35)%和(86.3±4.16)%，国外报道的最高成熟率为(85.7±4.1)%；第二部分对猪卵母细胞孤雌激活的化学方法进行了研究，确立了化学激活猪卵母细胞的两种最佳方法：1将成熟的去卵丘颗粒细胞的MII期卵母细胞用20μmol/Lionomycin作用30min，再将卵母细胞培养于含5μg/mlCB和5mM/L 6-DMAP(6-二甲基氨基嘌呤)的NCSU-23培养液中，卵裂率和桑囊胚发育率达到(76.7±7.6)%和(37.1±6.4)%2将成熟的去卵丘颗粒细胞的MII期卵母细胞在200μM/L的Thimerosal中处理20min，再与8mM的DTT共孵育30min，卵裂率和桑/囊胚形成率为(81.0±2.8)%和(39.6±2.7)%；第三部分对孤雌激活胚胎的培养条件进行了研究，确立了一种最佳的胚胎培养条件：在SOF简单培养基中添加颗粒细胞进行前3天的培养，然后转入添加胎牛血清的NCSU—23培养基并和输卵管上皮细胞进行后期的培养，其桑椹胚和囊胚的发育率为(59.5±3.2)%；第四部分研究了IGF-I

In the present study, we collected cumulus cells oocyte complex from ovaries of two different strain mice. The cumulusenclosed oocytes were cultured for 6 h in MEM supplemented with growth factor and FSH. The meiotic maturation of these oocytes has progressed to pro-metaphse Ⅰ stage and the condensed chromosomes are visible under DIC microscope, metaphase Ⅰ spindle even can be detected under Polscope. The metaphase Ⅰ spindles of oocytes were exchanged under such microscopes. After electric stimuli, 91. 6% and 91. 6% karyoplasts-cytoplasm pairs were fused respectively. The resulting oocytes were cultured further in MEM and over 80% of oocytes released the first polar body. 79% and 77% of oocytes formed two pronuclei after in vitro fertilization and the embryos were cultured in KSOM supplemented with amino acids. Over 60% of embryos developed to blastocyst stage.

在本研究中我们在取得两种不同品系小鼠的卵丘卵母细胞复合体后，先将卵丘卵母细胞复合体置于含有多种生长因子和激素的MEM培养液中培养6小时，此时卵母细胞已进入第一次减数分裂的前中期，并且在DIC倒置显微镜下可以看到浓缩的染色体，用Polscope可以发现明显的纺锤体，借助这种显微镜通过显微操作将两种不同品系小鼠来源的卵母细胞的MI纺锤体进行互换，经过三次直流电脉冲作用后，分别有91.6％的胞质—MI核质体对融合，经过进一步的培养后，超过80％的重组卵母细胞排出第一极体，体外受精后分别有79％和77％的重组卵形成双原核，受精后的胚胎在KSOM胚胎培养液中体外培养4天后，超过60％的胚胎发育至囊胚。

As she looked at Warrington's manly face, and dark, melancholy eyes, she had settled in her mind that he must have been the victim of an unhappy attachment.

每逢看到沃林顿那刚毅的脸，那乌黑、忧郁的眼睛，她便会相信，他一定作过不幸的爱情的受害者。

Maybe they'll disappear into a pothole.

也许他们将在壶穴里消失