摘要
本研究探讨了不同化学激活方法和不同电激活参数对猪体外成熟卵母细胞早期孤雌发育的影响。利用屠宰场猪卵巢卵母细胞，在体外成熟培养44～48h后，将核成熟卯母细胞分别用：（1）不同化学激活方法[①10％乙醇＋10μg／mL放线菌酮；②2．5mmol／L氯化锶＋10μg／mL放线菌酮；③5μmol／L离子霉素＋2．5mmol／L 6-二甲氨基嘌呤（6-DMAP）；④200μmol／L硫柳汞＋8mmol／L二硫苏糖醇进行激活处理；⑤对照组——不用任何激活剂（试验1）]；（2）用不同电场强度和脉冲时程进行电激活（试验2），之后放入胚胎培养液中进行体外培养7d。研究结果显示：（1）4种不同化学激活方法处理卵子的1原核形成率、2原核形成率和原核形成率都显著比对照组的高（P〈0．05），其中离子霉素＋6-DMAP组的2原核形成率和总原核形成率最高，分别为（23．1±3．5）％和（65．2±3．5）％，显著比其他处理组的都高（P〈0．05）；（2）4种不同化学激活方法处理组的囊胚率都比对照组的高（P〈0．01），其中离子霉素＋6-DMAP组的卯裂率及囊胚率最高，分别为（46．6±18．5）9／5和（5．6±4．2）％；（3）本研究所用的4种不同化学激活方法对猪4-细胞孤雌胚SDS—PAGE电泳的蛋白质表达图谱没有显著影响；（4）电场强度为1．7kV／cm、脉冲时程为50和70μs时猪体外成熟卯母细胞激活效果最好，卵裂率、囊胚率、囊胚细胞数分别为：（77．4±9．7）％，（12．4±3．7）％，17．6±5．9和（75．1±10．6）％，（12．3±2．6）％，19．1±8．1。以上结果说明：（1）本研究所用的4种化学激活方法均能有效激活猪体外成熟卵母细胞，其中离子霉素＋6-DMAP的激活效果最理想；（2）在本实验室条件下，采用1．7kV／cm的电场强度、50～70μs的脉冲时程均能有效的激活猪体外成熟卵母细胞；（3）本研究所用的4种不同化学激活方法对猪4-细胞孤雌胚SDS—PAGE电泳的蛋白质表达图谱没有显著影响。
The objectives were to investigate the effect of different chemical activation methods and electro-activation parameters of porcine oocytes matured and parthenogenetically activated in vitro. Porcine ooeytes aspircted from local abattoir ovaries were matured in vitro for 44～48 h. Oocytes with the first polar body were activated by ： （ 1 ） Different chemical methods [①10% ethanol ＋ 10 μg/mL cycloheximide （CHX） ;② 2. 5 mmol/L SrCl2＋10 μg/mL CHX; ③ 5 μmol/L Ionomycin＋2.5 mmol/L 6-dimethylaminopurine （DMAP） ;④ 200 μmol/L Thimerosal＋8 mmol/L dithiothreitol （DTT）; ⑤The control-no artificial activation） （Expt. 1）; （2） Different pulse strength and duration （ Expt . 2 ） , and then were cultured for 1 6 8 h in embryo culture medium . The results indicated ： （ 1 ） The rates of pronuclear formation of all treatment groups were higher than that of the control （P〈0. 05）, and the pronuclear formation rate of Ionomycin＋6-DMAP group which was the highest was significantly higher （P〈0.05） than those of other groups; （2） The blastocyst rates of all treatment groups were higher （P〈0.01） than that of the control; （3） The highest rates of cleavage and blastocyst were obtained in ionomycin＋6-DMAP group; （4） The SDS PAGE protein expression profiles of porcine 4-cell parthenogenetic embryos obtained from oocytes activated by the four chemical activation methods were not significantly different; （5） The efficiency of electro- activation of porcine oocytes matured in vitro was the best when the pulse strength was 1.7 kV/cm and the pulse duration was 50 μs or 70 μs. It was concluded that： （1） All of the chemical activation protocols used in this study significantly enhanced the blastocyst development of porcine oocytes, and the combination of ionomyein and 6-DMAP was the ideal artificial activation method for porcine oocytes; （2） The SDS- PAGE protein expression profiles of porcine 4 - cell parthenogenetic embryos obtained from oocytes activated by the four chemical activation methods were not significantly different ; （ 3 ） At our laboratory , the optimal electric stimulation parameter for porcine oocytes matured in vitro was 1.7 kV/cm pulse strength and 50 μs or 70 μs pulse duration.
出处
《广西农业生物科学》
CSCD
2006年第B09期184-185,共2页
Journal of Guangxi Agricultural and Biological Science
基金
广西亚热带生物资源保护利用重点实验室-省部共建国家重点实验室培育基地开放课题(SB0505)
广西科学基金项目(桂科青0640002和桂科青0447004)
广西科学研究与技术开发计划项目(桂科攻0537022-01和桂科攻0330004-14)
广州市华粤行仪器有限公司南宁办事处资助计划内横向项目
关键词
猪
卵母细胞
体外成熟
孤雌发育
胚胎培养
porcine
oocytes
in vitro maturation
parthenogenetic development
embryos culture
