Evaluation of Improvement and Optimization for Cryopreservation Technology of Chicken Semen with Glycerol
[Objective]The aim of this study was to improve and optimize the simple cryopreservation technique for glycerol straws in chicken semen.[Method]A single factor design experiment was conducted to compare the motility,viability,and abnormality rate of frozen semen using 6％cryopreservation(3,6 and 9 months)and freshly prepared glycerol protective solution.The one-step freezing rate was optimized using liquid nitrogen surface fumigation method.The motility,viability and abnormality rate of sperm,fertilization rate and hatching rate of breeding eggs after freezing and glycerol removal were tested in frozen semen prepared by the optimized one-step freezing and the two-step freezing developed.Using a one-step freezing method to freeze semen from different breeds of roosters,and applying different insemination cycles to hens of different ages,the effect of insemination was evaluated through insemination experiments.[Result]① There was no significant differences in motility,viability,or abnormality rate of semen preserved in the newly-prepared glycerol cryoprotectant or the preserved glycerol cryoprotectant for 3,6 and 9 months(P＞0.05).② The optimal freezing procedure of the one-step freezing was that the straw was set 3 cm above the liquid nitrogen surface for 3 min to achieve sperm motility above 60％in all replicates,and above 65％in 65％replicates.③The motility and viability of sperm were 62.84％and 76.00％in one-step freezing,which were extremely significantly higher than those in two-step freezing(58.17％and 68.62％)(P＜0.01).The sperm abnormality rate was no significantly difference between the two groups(P＞0.05).There was no significant difference in the average sperm motility,average fertilization rate,and average hatching rate of breeding eggs between the one-step and two-step frozen semen centrifugation before and after glycerol removal(P＞0.05).The highest daily fertilization rates were 68.34％and 80.00％,respectively.④ There was no significant difference in the fertilization rate and hatching rate of the eggs prepared by one-step freezing method between Heifeng chicken and Xiangdong chicken after frozen sperm insemination(P＞0.05).The fertilization rate of eggs after insemination to 30 week old hens was extremely significantly higher than that of 68 week old hens(P＜0.01).The fertilization rate of eggs in the 3 round 4 insemination mode was significantly extremely higher than that in the 1 round 2 insemination mode(P＜0.01).[Conclusion]It could achieve a better effect of insemination by using-20 ℃ frozen glycerol as cryoprotectant and one-step cryopreservation technology.Attention should be paid to the time of semen refrigeration,the age of the hen,and the number of insemination rounds during the insemination for a stable and better effect.
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