The Study On Freezing For KUNMING Wolfhound Sperm

The kunming wolfhound were regarded as the study objects.Have done the study of diferent centrifugal rate,the semen dilutes screening and optimization,equilibration method,Frozen dosage form and frozen methods,artificial fertilization of frozen semen ect. The result were as follow:1.Effects of diferent centrifugal rate on sperm motility after frozen-thawed:the 5 dogs's fresh semen were carded on centrifugal with Or/min(contrast group),1000 r/ min,1500 r/min,2000 r/rain,2500 r/min,3000 r/min,3500 r/min,4000 r/min.the 1st dilution was selected which replaced and freezed with ultra-low temperature after the 1st dilution was washed repeatedly.After the 1st dilution was unfreezed and measure the sperm motility.The result proved:the 1500 r/rain group's sperm motility is 43.5±2.43,which compared to the 2000 r/min group is not prominent(P＞0.05).compared to other groups,the 1500 r/min and 2000 r/min two groups are significant difference(P＜0.05). The result showed that the effect of centrifugal at 1500r/min is the best,acrosome intact rate is obviously higher than other groups(P＜0.05),The sperm abnormal rate is the lowest (24.5±1.87),survival index is the highest(28.2±1.24) among the groups.Unfreezed under 4℃,sperm's survival time amounts to 96 hours.2.The dilution screening and optimization:TheⅠ,Ⅱ,Ⅲdilution groups that selected were made pellet semen.After unfreezed,the result indicated:ⅠGroup average sperm motility superior toⅡandⅢgroups,but there was no difference betweenⅠandⅡgroups (P＜0.05).TheⅠandⅡgroups compared withⅢgroup,there were significant difference (P＜0.05).TheⅠgroup's acrosome intact rate was superior to others and sperm abnormal rate was lower than others(P＜0.05).In the course of semen dilution optimize,Ⅰadded to amount of V_B12,C_OV_B or V_C separately can improve the freeze-unfreeze sperm motility and can protect acrosome intact and can reduce sperm abnormal rate,but compared with contrast group,there was no difference(P＜0.05).3.The study of semen equilibration methods:The result showed:The dilution was equilibrated by wrapped up by 16 layers,8 layers of gauzes and bathed in the 100ml water,200ml water,300ml water.The results,it showed:The dilution was equilibrated by wrapped up with 16 layers and keeped in the refrigerator at 4℃for 3.5-4.0 hours.In this way,the unfreezed-semen motility was 43.23%,survival index was 27.22±2.45 and acrosome intact rate was 77.59%.Compared with bathed in the 200ml water,it was no difference(P＞0.05),but it was significant difference than ather groups(P＜0.05).4.The study of frozen dosage form and frozen methods:the metheds of bombing stifling,liquid nitrogen aluminium lunch-box,thin tube bombing stifling and Ann's bottle stifling for frozen were used in the frozen-unfreeze dilution.The result showed:the sperm motility with bombing stifling is 43.48±2.07,it was better than others(P＜0.05).With the procedure of refrigerationⅠ,Ⅱ,Ⅲ,made thin tube frozen semen.After unfreezed,the sperm motility is 32.08±3.16,which superior toⅡ(25.26±0.92) andⅢ(14.57±2.09)(P＜0.05).But the bombing stifling frozen semen is the best than others(P＜0.05).5.The study of unfreeze methods:With dry unfreeze and wet unfreeze the pellet semen,the result showed:On the condition of bathing at 50～55℃,the pellet semen unfreezed,which sperm average motility is 42.30%.It is better than wet unfreeze pellet semen(P＞0.05).With dry unfreeze,the pellet semen's acrosome intactrate(67.01±2.20) is better than others(p＜0.05),but which sperm abnormal rate is significant difference(p＜0.05).6.The study of artificial fertilization of frozen semen:Divide 10 female dogs into the frozen semen group(6),fresh semen group(4) at random.With the same male dog semen,which was filled in the uterus every 24 hours for 1 times and for 3 times.The result showed:The impregnating rate of frozen semen is 50.00%,the fresh semen impregnating rate is 75.00%.