| Semen cryopreservation technology solved the problem of the long-term preservation of semen,so that semen is no longer limited by seasons,regions and the life of breeding animals,and accelerates the progress of animal husbandry breed improvement and cultivation.With the development of modern animal husbandry,the demand for frozen semen in artificial insemination is increasing,and the quality requirements for frozen semen are getting higher and higher.In order to improve the quality of frozen semen of sheep,this study took Turpan black sheep ram as the experimental object,and added different concentrations(0,5,15,25,35 μg/m L)of(proanthocyanidins,PC)to three different semen freezing dilutions respectively.Tris-anhydrous glucose-anhydrous citric acid is liquid I,Tris-glucose-citric acid is liquid II,and Optidyl dilution is used as control liquid III.By detecting frozen-thawed sperm motility rate,motility parameters,deformity rate,CAT activity,GSH-Px activity,T-AOC,plasma membrane integrity rate,acrosome integrity rate,mitochondrial membrane potential,sperm DNA fragmentation and artificial insemination fertilization rate,The formula of diluention with the best effect on cryopreservation of Turpan black sheep semen and the best concentration of PC were selected out.The main results obtained in this study are as follows:(1)Before freezing,the 15 μg/m L PC group of liquid II had the highest viability rate of88.52%,which was higher than that of the 0 and 5 μg/m L PC groups of liquid I(P < 0.05),and was higher than that of the 15 μg/m L PC group of liquid I(P < 0.01);After freezing,the 15 μg/m L PC group of liquid I had the highest motility rate of 73.307%,which was higher than that in the 15μg/m L PC group in liquid III(P < 0.05).The motility rate of the 5 μg/m L PC group in liquid I was69.450%,which was not different from the 15 μg/m L PC group in liquid II and the 25 μg/m L PC group in liquid III(P > 0.05).There was no significant difference in the deformity rate among the groups before freezing(P > 0.05).After freezing,the deformity rate in the 15 μg/m L PC group in liquid I was the lowest at 9.845%,which was not significantly different from the 15,25 μg/m L PC group of liquid III,and the 5,25 μg/m L PC group in liquid I(P > 0.05).After freezing,the kenematic parameters including VCL,VSL,LIN,VAP,ALH,WOB of the 15 μg/m L PC group in liquid I were higher than those of other groups(P < 0.01);There was no significant difference in kinematic parameters such as VSL,VCL,ALH,and WOB in the 25 μg/m L PC group in liquid II(P > 0.05).Based on the above results,the routine test results of semen quality after thawing were better in the 5 and 15 μg/m L PC group in liquid I,15 μg/m L PC group in liquid II,15 and 25μg/m L PC group in liquid III,the activity of antioxidant enzymes and sperm microstructure can be further detected to determine the optimal freezing formulation and optimal PC addition concentration.(2)The CAT activity of the 15 μg/m L PC group in liquid I was higher than that in the 15μg/m L PC group in liquid II(P < 0.05),but not significantly different from other groups(P >0.05).The GSH-Px activity of the 15 μg/m L PC group in liquid I was significantly higher than that in the 5 μg/m L PC group in liquid I(P <0.05),and extremely significantly higher than that in the other three groups(P <0.01).The content of T-AOC in the 15 μg/m L PC group of liquid I was the highest,and there was no significant difference with that of the 25 μg/m L PC group in liquid III(P > 0.05).The integrity rate of plasma membrane in the 15 μg/m L PC group in liquid I was higher than that of the 5 μg/m L PC group in liquid I and the 15 μg/m L PC group in liquid II(P < 0.01).The acrosome integrity rate of 15 μg/m L group in liquid I was higher than that of 15μg/m L PC group in liquid II and 25 μg/m L PC group in liquid III(P < 0.01).The ratio of mitochondrial membrane potential in the15 μg/m L PC group of liquid I was higher than that of 15μg/m L PC group in liquid II(P < 0.05),but not significantly different from the other three groups(P > 0.05).The sperm DNA fragmentation rate in the 15 μg/m L PC group in liquid I was significantly lower than that of 15 μg/m L PC group in liquid II and the 5 μg/m L PC group in liquid I(P < 0.05),and there was no significant difference between the other two groups(P >0.05);After artificial insemination,the fertilization rate in the fresh semen group was significantly higher than that in the frozen semen groups,but among the frozen semen groups,the fertilization rate of 15μg/m L PC group in liquid I was higher than that in the other 4 groups(P < 0.05).In conclusion,the formula of adding 15 μg/m L PC to Tris-anhydrous glucose-anhydrous citric acid freezing dilution significantly improved the cryopreservation effect of Turpan black sheep semen. |
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