关键词: 哈茨木霉, 哈茨木霉, 对峙培养, 拮抗, 抗药性

Abstract: 【Objective】 In order to understanding effects of different treatment methods of frozen semen and addition of caffeine on IVF in bovine, 【Methods】 frozen-semen of bovine was treated by using Swimming-up method, Percoll method and Washing method, capacitated by adding 2.5mM or 5.0mM of caffeine in capacitation medium(BO medium+10μg/ml heparin sodium) and pre-incubated 10min or 30min to observe respective effect on the in vitro development ability of fertilized eggs. 【Results】 The results showed that (1)there was not any significant differences among cleavage rate(51.58%,52.83%,53.20%)(P＞0.05) of fertilized eggs treated by frozen-semen from Washing method ,Swimming-up method and Percoll method，there was also not any significant differences on the in vitro development rate of morula or blastocyst between treatment of Percoll method(16.66%,10.18%) and Swimming-up method(17.85%,16.07%),and between treatment of Percoll method and of Washing method(8.77%, 7.01%)(p＞0.05).But it can be seen from experimental results that in vitro development of morula and blastocyst of treatment of Swimming-up method had some enhance compared with that of treatment of Percoll method, there was significant differences of in vitro development rate of morula or blastocyst between treatment of Swimming-up method and of Washing method(P＜0.05).(2)Addition of caffeine in fertilization medium can promote activity of sperm. Cleavage rate or development rate of morula or blastocyst in the presence of caffeine were higher than that of in the absence of caffeine, there was best effect of capacitation on sperm（Cleavage rate is 68.59%，blastocyst rate is 28.50%）.【Conclusion】This indicated that, compared these 3 methods, Swimming-up method perhaps is economical and simple method of treatment methods of frozen-semen, it can be applied for IVP of embryo of bovine; This also showed that higher concentration of caffeine can maintain strong activity of sperm at comparative long period of time.