Research On Preservation Of Goat Semen

Goat semen preservation technology can improve the use efficiency of the excellent ram,accelerate the pace of improced varieties.It can facilitate long-distance transportation of semen,reduce the risk of disease transmission.But the goat semen low temperature storage and cryopreservation technology is not mature,we need to further optimize.In this study,we chose Anhui white goat as the object,carried a systematic study on the effect of goat semen low temperature storage and cryopreservation,the contents are as follows:One,the effect of pH on the goat semen low temperature storage.Acquisition of the mixed semen was divided into five parts,and semen was diluted by pH 6.15,6.25,6.35,6.45 and 6.80 extenders,stored them in 5 degrees refrigerator,checked the semen quality once every two days.Result: Saved 2 days,Sperm motility of pH 6.35 group was significantly higher than that of other groups(P<0.05);on the second,sixth,eighth,tenth and twelfth days,the percentages of integrity of membrane and acrosome of pH 6.35 extender were significantly higher than that of other groups(P<0.05);Used pH 6.35 extender to store semen,on the fourth,tenth and twelth days,before insemination,raised the buffer pH to 7.2,the motility was significantly higher than that of the control group(did note raise pH)(P<0.05).Two,the effect of osmotic pressure on the goat semen low temperature storage.Acquisition of the mixed semen was divided into four parts,and semen was diluted by extenders of 330 m Osm,350 mOsm,375mOsm and 400 mOsm osmotic pressure respectively,stored them in 5 degrees refrigerator,checked the semen quality once every two days.Result: 8 days later,the percentages of sperm motility of 375 mOsm group was significantly higher than that of other groups(P<0.05),6 days later,the percentages of integrity of membrane and acrosome of 375 mOsm group were significantly higher than that of other groups(P<0.05).Three,used optimized method to store goat semen,used the semen that saved for3,6,9 days and fresh semen to inseminate the oestrus ewes.Result: the non-return oestrus rates of the semen that saved for 3,6 days were more than 70%,met the request of insemination.Though the non-return oestrus rate of the semen that saved for 9 day was significantly lower than that on the third day,but it also reached63.11%.Four,did a series of tests,optimizate goat semen cryoperservation: Choice 4~6years old rams,basic extender was prepared consisting of tris 3.07g/100 mL,fructose1.26g/100 mL,citric acid 1.64g/100 mL,egg yolk 15%(V/V),diluted the semen in 2steps,finally,the sperm density was 2.4×108/mL,cooling equilibrium time was 4h,the final concentration of glycerol was 5%(V/V).When glycerol was added,the second equilibrium time was 30 min.The fumigation height above the liquid nitrogen was 4cm.Fumigating time was 10 min,then plunged the pellet into the liquid nitrogen,and thaw in 37 degrees for 30 seconds.Five,the effect of using the substitution of cholestero(CHL)for egg yolk(EY)to store goat sperm.Acquisition of the mixed semen was divided into five parts,use 1g/L(group C1),2.5 g/L(group C2.5),5 g/L(group C5),10.0g/L(group C10)CHL and 15% EY(control group)to dilute semen respectively and then frozen.After thawing,used sperm computer-assisted system(SCAS)to detecte sperm kinematic parameters,used hypoosmotic swelling test(Host)to detecte the membrane intact rate and used giemsa staining to detect the acrosome intact rate.Result: The post-thaw sperm motility,acrosome intact rate,curvilinear velocity(VCL),velocity of average path(VAP)of control group were significantly higher than that of CHL addition groups(P<0.05),the plasma membrane integrity rate,linearity(LIN),straightness(STR),wobble(WOB),amplitude of lateral head displacement(ALH)and beat-cross frequency(BCF)of control group had no significant difference with that of C2.5,C5,C10 and EY groups(P>0.05),but the plasma membrane integrity rate of control group was significantly higher than that of C1 group(P<0.05).Six,the effect of using the substitution of phosphatidylcholine(PC)for egg yolk(EY)to store goat sperm.Acquisition of the mixed semen were divided into five parts,used 10 g/L(group P10),15 g/L(group P15),20 g/L(group P20),25 g/L(group P25)PC and 15% EY(control group)to dilute the semen respectively and then frozen.Result: there was no significant differences between P15 group and EY group in motility,VCL,VAP,LIN,STR,WOB and BCF(P>0.05),the motility of P10,P20 and P25 was significantly lower than that of EY group(P<0.05).Seven,the change of cholesterol and phospholipids content in sperm membrane before and after freeze-thaw.Membrane phospholipid content of goat fresh semen was 559.5±20.23 nmol/109,the content of cholesterol was 320.0±16.7 nmol/109.Used basic extender(withou yolk)to dilute semen.The phospholipid content of post-thaw goat semen was 420.3±18.1 nmol/109,the content of cholesterol was 130.7±20.8nmol/109,the quality ratio of lost phospholipid and cholesterol in post-thaw sperm was 3.759:1.Eight,the effect of using the substitution of PC-CHL for egg yolk(EY)to store goat sperm.According to the results of test seven,the design of PC-CHL formula was as follows: basic extender+19g/L PC+1g/L CHL(P19+C1);basic extender+18g/L PC+2g/L CHL(P18+C2);basic extender+16g/L PC+4g/L CHL(P16+C4);basic extender+14g/L PC+6g/L CHL(P14+C6),basic extender+15% EY(EY).Result:the post-thaw motility,VCL,velocity of straight-line(VSL)and VAP of P19+C1 and P18 + C2 were not significantly different from the control group(P>0.05);the motility,VCL and VAP of P16+C4 group were significantly higher than that of the control group(P<0.05),the motility in P14+C6 was significantly higher than that in control group(P<0.05);After freezing-thawing,the phospholipid content of P16+C4group was significantly higher than that of the other groups(P<0.05),cholesterol content was lower than EY group,but the difference was not significant(P>0.05).Nine,used P16+ C4 extender and EY extender to freeze goat semen and use them for artificial insemination.Result: the non-return oestrus rate,conception rate and lambing rate of P16+C4 group were higher than that of EY group.Conclusion,1.Low temperature storage of goat semen in 5 degree,the optimal buffer pH is 6.35,the optimal osmotic pressure is 375 mOsm.Before insemination,improving semen pH to 7.2 can improve sperm motility.2.Adding 14g/L PC and 6g/L CHL at the same time can replace 15% egg yolk for goat semen cryopreservation.