【摘要】 卵巢卵母细胞培养于A、B、C和D4种成熟液内的成熟率分别为83.3％,8.3％,55.8％。和100％。在A、C和D中培养成熟的卵母细胞,体外受精率分别为44.4％,54.5％。和71.6％。授精卵中发育至2-细胞,4-细胞和8-细胞期者分别为32.4％,23.7％和14.0％。2-细胞期胚胎中发育到桑椹胚和囊胚者分别为56.8％和35.1％。移植鲜胚受体5头,妊娠2头,其中1头胚胎死亡,另1头流产;移植冻胚6头受体,其中1584号受体于1989年9月23日产一雄性牛犊,初生重36.5公斤。妊娠期为282天。更多还原

【Abstract】 The objective of the present study was to obtain calves from bovine follicular oocytes matured, fertilized, developed in vitro and frozen-thawed embryos. Ovaries were collected from local slaughterhouse and brought to laboratory within one hour, held at 30-38℃ in sterile saline, Oocytes were obtained from follicles ( 2-5 mm in diameter ) of ovaries by aspiration with a needle and liberated by follicle rupture. The average numbers of oocytes and useful oocytes collected from each ovary by follicle rupture (10.5 and 8.3) were more than those corresponding values by aspiration ( 2.3 and 2.0 ) . Those oocytes with a compact cumulus cells were cultured in a droplet of maturation medium under mineral oil for 22-24 hrs at 38.5℃ , 5% CO2 in air with maximum humidity. Four maturation media ( A,B,C and D ) were used and maturation rates were 83.3%, 8.3%, 55.8% and 100% respectively. Frozen bull semen was thawed at 35-37℃ and washed twice in BO medium ( Brackettand Oliphant, 1975) supplemented with 10 mM caffeine by centrifuga-tion at 350 g for 10 min. The sperm were then suspended in a 2 ml of medium (3.0×10~6 sperm/ml) and treated with 0.1 (μM of the iono-phore A 23187 for 1 min. A 50 ul aliquot of the sperm suspension was introduced into a microdrop of 50 ul BO medium supplemented with 20 mg bovine serum albumin/ml, containing 20-30 oocytes and then cultured for 6 hrs. Fertilization rates were 44.4%, 54.5% and 71.6% of oocytes matured in maturation medium A, G, and D respectively. For in vitro development, the inseminated oocytes were cultured for 7-9 days in 350 μl of TCM-199 plus 10% FCS. Of 207 oocytes, 67 ( 32.4% ) cleaved to the 2-cell stage and 49(23.7%) to 4-cell stage and 29(14%) to 8-cell stage. Development rates of 2-cell stage reached morulae and blastocyst were 56.8% and 35.1% respectively. Two of 5 recipients transferred by non-surgical with fresh embryos established pregnancies but did not give birth of calves as the result of death of early embryos in one recipient and of abortion in the other. One of 6 recipients transferred with frozen embryos was pregnancy and born a~healthy calf on Sep. 23,1989.更多还原