不同抗冻剂及初冻和解冻温度对鸡精液冷冻保存的影响
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摘要
【目的】研究对不同抗冻剂及初冻和解冻温度在鸡精液冷冻保存中的效果.【方法】以‘海兰褐’种公鸡为试验材料,利用液氮制作颗粒冷冻精液,研究不同抗冻剂及初冻和解冻温度对鸡精液冻后活率、复苏率及存活时间的影响,并筛选出最佳的抗冻剂种类、浓度和初冻、解冻温度组合.【结果】3种抗冻剂组合的冷冻效果优于单一及两种抗冻剂组合,且GL 8%+DMA 6%+EG 6%组合的冻后活率、精子复苏率最高,分别为0.46和57.50%,为最佳抗冻剂组合;在此基础上,筛选出最佳初冻和解冻温度组合为-140℃和50℃,其冻后活率和复苏率最高,分别达到0.57和71.40%,并在15～25℃条件下精子存活时间为160min;在鸡精液冷冻保存中快速冷冻要对应快速解冻,慢速冷冻要对应慢速解冻,且前者精液冷冻保存效果较好.【结论】筛选出了最佳的抗冻剂组合及初冻和解冻温度,发现快速冷冻和解冻的鸡精液保存效果较好.
【Objective】In this study,the cryoprotectant,start freezing temperature and thawing temperature were researched as the key factors to provide foundational data for optimizing chicken semen cryopreservation.【Method】The‘Hy-Line'breeder cocks were used as experimental materials,with pellet frozen semen made by liquid nitrogen to research the effect of the cryoprotectant,start freezing temperature and thawing temperature on frozen rate,sperm recovery rate and survival time and screen the best cryoprotectant and the combination of start freezing and thawing temperature.【Result】The freezing effect of three cryoprotectant combinations was superior to the single and two kinds of cryoprotectant,one group of GL(8%)+DMA(6%)+EG(6%)with highly frozen rate(0.46)and sperm recovery rate(57.50%)was the optimum cryoprotectant combination.On this basis,the best combination of start freezing and thawing temperature screened out were -140 ℃ and 50 ℃,the frozen rate and sperm recovery rate were 0.57 and 71.40%,and the survival time at 15～25 ℃ was up to 160 minutes.The rapid frozen should be corresponded to rapid thawing,and the slow frozen should be corresponded to slow thawing,and the former had bet-ter effect on chicken semen cryopreservation.【Conclusion】The optimum combinations of cryoprotectant and freezing and thawing temperature are screened,and the rapid frozen thawed shows better on chicken semen preservation.
【Objective】In this study,the cryoprotectant,start freezing temperature and thawing temperature were researched as the key factors to provide foundational data for optimizing chicken semen cryopreservation.【Method】The‘Hy-Line'breeder cocks were used as experimental materials,with pellet frozen semen made by liquid nitrogen to research the effect of the cryoprotectant,start freezing temperature and thawing temperature on frozen rate,sperm recovery rate and survival time and screen the best cryoprotectant and the combination of start freezing and thawing temperature.【Result】The freezing effect of three cryoprotectant combinations was superior to the single and two kinds of cryoprotectant,one group of GL(8%)+DMA(6%)+EG(6%)with highly frozen rate(0.46)and sperm recovery rate(57.50%)was the optimum cryoprotectant combination.On this basis,the best combination of start freezing and thawing temperature screened out were -140 ℃ and 50 ℃,the frozen rate and sperm recovery rate were 0.57 and 71.40%,and the survival time at 15～25 ℃ was up to 160 minutes.The rapid frozen should be corresponded to rapid thawing,and the slow frozen should be corresponded to slow thawing,and the former had bet-ter effect on chicken semen cryopreservation.【Conclusion】The optimum combinations of cryoprotectant and freezing and thawing temperature are screened,and the rapid frozen thawed shows better on chicken semen preservation.
引文
[1]BLESBOIS E.Current status in avian semen cryopreservation[J].World Poultry Science Journal,2007,63(2):213-222.
[2]LAKE P E.The history and future of the cryopreservation of avian germ plasm[J].Poultry Science,1986,65:1-15.
[3]GLIOZZI T M,ZANIBONI L,CEROLINI S.DNA fragmentation in chicken spermatozoa during cryopreservation[J].Theriogenology,2011,75(9):1613-22.
[4]CHENG C Y,CHEN P R,CHEN C J,et al.Differential protein expression in chicken spermatozoa before and after freezing-thawing treatment[J].Animal Reproduction Science,2015,152:99-107.
[5]DONOGHUE A M,WISHART G J,Storage of poultry semen[J].Animal Reproduction Science,2000,62:213-232.
[6]张凯,吴海云,张立昌,等.抗冻剂和冷冻速率对蛋用种公鸡精液冷冻保存效果的影响[J].中国农学通报,2014,30(2):13-17.
[7]黄素红.鸡精液冷冻保存技术研究[D].兰州:甘肃农业大学,2007.
[8]IAFFALDANO N,ROMAGNOLI L,MANCHISI A,et al.Cryopreservation of turkey semen by the pellet method:effects of variables such as the extender,cryoprotectant concentration,cooling time and warming temperature on sperm quality determined through principal components analysis[J].Theriogenology,2011,76(5):794-801.
[9]GROSSFELD R,SIEG B,STRUCKMANN C,et al.New aspects of boar semen freezing strategies[J].Theriogenology,2008,70(8):1225-1233.
[10]张兆旺,刘丽霞.稀释液溶质、渗透压、pH值和缓冲指数与低温保存鸡精液效果的关系[J].中国家禽,2004,26(21):13-16.
[11]赵建军,张伟,张兆旺.冷冻稀释液渗透压、抗冻剂和平衡时间对鸡颗粒冻精冻后活率的影响[J].甘肃农业大学学报,2006,41(5):14-17.
[12]王世银,张伟,张兆旺,等.抗冻剂对鸡精液冷冻效果的影响[J].江苏农业科学,2011,39(3):260-262.
[13]王峰.动物繁殖学[M].北京:中国农业大学出版社,2012.
[14]徐日福,王红军,张忠诚.鸡精液最佳冷冻时间和解冻温度的确定[J].江西畜牧兽医杂志,1997(4):13-14.
[15]张坚中,陈元明,邵桂芝.绵羊精液冷冻保存技术[J].农业科技通讯,1984(10):26.
[16]郭年藩.前苏联家禽人工授精技术研究的回顾(上)[J].国外畜牧兽医,1997,24(4):38-42.
[17]BUSS E G.Cryopreservation of rooster sperm[J].Poultry Science,1993,72(5):944-54.
[18]TAI J J L,CHEN J C,WU K C,et al.Cryopreservation of gander semen[J].British Poultry Science,2001,42(3):384-388.
[19]HAN X F,NIU Z Y,YANG C S,et al.Effects of diluents,cryoprotectants,equilibration time and thawing temperature on cryopreservation of duck semen[J].International Journal of Poultry Science,2005,4(4):197-201.
[20]王红军,金凤英.混合冷冻保护剂在鸡精液冷冻中的作用分析[J].中国畜牧杂志,1998(1):31-32.
[21]徐日福,王红军,张忠诚.种鸡精液冷冻保护剂的确定[J].黑龙江畜牧兽医,1996(11):16-18.
[22]王元兴,郎介金.动物繁殖学[M].南京:江苏科学技术出版社,1997.
[23]BELLAGAMBA F,CAVALCHINI S C L G.Cryopreservation of poultry semen:a review[J].Worlds Poultry Science Journal,1993,49(2):157-166.
[24]潘瑞.渗透压、初冻温度及保存期对鸡精子冻后活率的影响[D].兰州:甘肃农业大学,2008.
[25]王肖克,杨健.影响家畜冷冻精液质量的因素[J].内蒙古畜牧科学,2002(6):20-21.
[26]卜书海,孙志杰,李青旺,等.不同解冻方法对奶牛冻精活力的影响[J].西北农林科技大学学报(自然科学版),2002(5):40-43.
[27]李世英,李焕玲,胡松庭,等.家禽精液冷冻保存技术的研究进展[J].中国家禽,1995(4):24-26.
[28]黄琳,孙鏖,朱立军,等.黑丝羽乌骨鸡精液DMA颗粒冷冻技术研究[J].中国畜牧兽医,2017(2):447-455.
[29]邓溯顺,燕海峰,易康乐,等.家禽精液冷冻保存技术研究进展[J].家禽科学,2012(5):45-48.
[30]KOWALCZYK A.The effect of cryopreservation process on morphology and fertilising ability of Japanese quail(Coturnix japonica)spermatozoa[J].Cryo Letters,2008,29(3):199.
[2]LAKE P E.The history and future of the cryopreservation of avian germ plasm[J].Poultry Science,1986,65:1-15.
[3]GLIOZZI T M,ZANIBONI L,CEROLINI S.DNA fragmentation in chicken spermatozoa during cryopreservation[J].Theriogenology,2011,75(9):1613-22.
[4]CHENG C Y,CHEN P R,CHEN C J,et al.Differential protein expression in chicken spermatozoa before and after freezing-thawing treatment[J].Animal Reproduction Science,2015,152:99-107.
[5]DONOGHUE A M,WISHART G J,Storage of poultry semen[J].Animal Reproduction Science,2000,62:213-232.
[6]张凯,吴海云,张立昌,等.抗冻剂和冷冻速率对蛋用种公鸡精液冷冻保存效果的影响[J].中国农学通报,2014,30(2):13-17.
[7]黄素红.鸡精液冷冻保存技术研究[D].兰州:甘肃农业大学,2007.
[8]IAFFALDANO N,ROMAGNOLI L,MANCHISI A,et al.Cryopreservation of turkey semen by the pellet method:effects of variables such as the extender,cryoprotectant concentration,cooling time and warming temperature on sperm quality determined through principal components analysis[J].Theriogenology,2011,76(5):794-801.
[9]GROSSFELD R,SIEG B,STRUCKMANN C,et al.New aspects of boar semen freezing strategies[J].Theriogenology,2008,70(8):1225-1233.
[10]张兆旺,刘丽霞.稀释液溶质、渗透压、pH值和缓冲指数与低温保存鸡精液效果的关系[J].中国家禽,2004,26(21):13-16.
[11]赵建军,张伟,张兆旺.冷冻稀释液渗透压、抗冻剂和平衡时间对鸡颗粒冻精冻后活率的影响[J].甘肃农业大学学报,2006,41(5):14-17.
[12]王世银,张伟,张兆旺,等.抗冻剂对鸡精液冷冻效果的影响[J].江苏农业科学,2011,39(3):260-262.
[13]王峰.动物繁殖学[M].北京:中国农业大学出版社,2012.
[14]徐日福,王红军,张忠诚.鸡精液最佳冷冻时间和解冻温度的确定[J].江西畜牧兽医杂志,1997(4):13-14.
[15]张坚中,陈元明,邵桂芝.绵羊精液冷冻保存技术[J].农业科技通讯,1984(10):26.
[16]郭年藩.前苏联家禽人工授精技术研究的回顾(上)[J].国外畜牧兽医,1997,24(4):38-42.
[17]BUSS E G.Cryopreservation of rooster sperm[J].Poultry Science,1993,72(5):944-54.
[18]TAI J J L,CHEN J C,WU K C,et al.Cryopreservation of gander semen[J].British Poultry Science,2001,42(3):384-388.
[19]HAN X F,NIU Z Y,YANG C S,et al.Effects of diluents,cryoprotectants,equilibration time and thawing temperature on cryopreservation of duck semen[J].International Journal of Poultry Science,2005,4(4):197-201.
[20]王红军,金凤英.混合冷冻保护剂在鸡精液冷冻中的作用分析[J].中国畜牧杂志,1998(1):31-32.
[21]徐日福,王红军,张忠诚.种鸡精液冷冻保护剂的确定[J].黑龙江畜牧兽医,1996(11):16-18.
[22]王元兴,郎介金.动物繁殖学[M].南京:江苏科学技术出版社,1997.
[23]BELLAGAMBA F,CAVALCHINI S C L G.Cryopreservation of poultry semen:a review[J].Worlds Poultry Science Journal,1993,49(2):157-166.
[24]潘瑞.渗透压、初冻温度及保存期对鸡精子冻后活率的影响[D].兰州:甘肃农业大学,2008.
[25]王肖克,杨健.影响家畜冷冻精液质量的因素[J].内蒙古畜牧科学,2002(6):20-21.
[26]卜书海,孙志杰,李青旺,等.不同解冻方法对奶牛冻精活力的影响[J].西北农林科技大学学报(自然科学版),2002(5):40-43.
[27]李世英,李焕玲,胡松庭,等.家禽精液冷冻保存技术的研究进展[J].中国家禽,1995(4):24-26.
[28]黄琳,孙鏖,朱立军,等.黑丝羽乌骨鸡精液DMA颗粒冷冻技术研究[J].中国畜牧兽医,2017(2):447-455.
[29]邓溯顺,燕海峰,易康乐,等.家禽精液冷冻保存技术研究进展[J].家禽科学,2012(5):45-48.
[30]KOWALCZYK A.The effect of cryopreservation process on morphology and fertilising ability of Japanese quail(Coturnix japonica)spermatozoa[J].Cryo Letters,2008,29(3):199.