幼龄羊的周龄及卵母细胞体外培养体系对JIVET技术的影响
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摘要
为了探讨周龄及卵母细胞体外培养体系对幼龄羊胚胎体外生产技术(JIVET)的影响,试验对10只5~10周龄杜寒杂种羔羊进行激素诱导,获取卵母细胞,分别在基础成熟液和含200μmol/L半胱氨酸的成熟液里成熟,再分别在含2%发情羊血清(ESS)的Ⅰ受精液和含3 mg/m L牛血清白蛋白(BSA)的Ⅱ受精液里受精,测定不同周龄羔羊的采卵数量、卵母细胞成熟率和卵裂率。结果表明:5,7,8,10周龄羔羊获得卵母细胞数分别为(111.00±16.97)枚/只、(139.50±28.99)枚/只、(108.50±17.68)枚/只、(42.00±11.31)枚/只,5,7,8周龄羔羊显著高于10周龄羔羊(P<0.05),而5,7,8周龄羔羊之间差异不显著(P>0.05);试验成熟液获得的卵母细胞成熟率比基础成熟液高3.64百分点;Ⅰ受精液获得的卵母细胞卵裂率极显著高于Ⅱ受精液(P<0.01)。说明选择5~8周龄羔羊、卵母细胞体外成熟液添加一定量半胱氨酸和体外受精液含一定量ESS可以改善JIVET体系。
The aim of the present study was to investigate the effects of week-old and in vitro culture system of oocytes on in vitro production technology of juvenile sheep embryos( JIVET). 10 Dorper-Small tailed hybrid lambs aged 5-10 weeks were induced by hormone to obtain oocytes. The oocytes were matured in the basal maturation solution and the experimental maturation solution containing 200 μmol/L cysteine,respectively. Then the oocytes were fertilized in fertilization medium Ⅰ containing 2% estrus sheep serum( ESS) and fertilization medium Ⅱ containing 3 mg/m L bull serum albumin( BSA). Finally,the number of eggs,oocyte maturation rate and cleavage rate of the lambs at different week-old were determined. The results showed that the number of oocytes obtained from the Dorper-Small tailed hybrid lambs at 5,7,8 and10-week-old were( 111. 00±16. 97) per lamb,( 139. 50 ± 28. 99) per lamb,( 108. 50 ± 17. 68) per lamb and( 42. 00 ± 11. 31) per lamb,respectively. The number of oocytes obtained oocytes from 5,7 and 8-week-old lambs was significantly higher than that from 10-week-old lambs( P< 0. 05),but there was no significant difference among 5,7 and 8-week-old lambs( P> 0. 05). The maturation rate of oocytes obtained from the test maturation solution was 3. 64 percentage point higher than that from the basic mature solution. The cleavage rate of oocytes from fertilization medium Ⅰ was significantly higher than that from fertilization medium Ⅱ( P< 0. 01). The results indicated that selection of lambs aged 5-8 weeks,oocytes in vitro maturation solution containing a certain amount of cysteine and fertilization medium containing a certain amount of ESS could improve the JIVET system.
The aim of the present study was to investigate the effects of week-old and in vitro culture system of oocytes on in vitro production technology of juvenile sheep embryos( JIVET). 10 Dorper-Small tailed hybrid lambs aged 5-10 weeks were induced by hormone to obtain oocytes. The oocytes were matured in the basal maturation solution and the experimental maturation solution containing 200 μmol/L cysteine,respectively. Then the oocytes were fertilized in fertilization medium Ⅰ containing 2% estrus sheep serum( ESS) and fertilization medium Ⅱ containing 3 mg/m L bull serum albumin( BSA). Finally,the number of eggs,oocyte maturation rate and cleavage rate of the lambs at different week-old were determined. The results showed that the number of oocytes obtained from the Dorper-Small tailed hybrid lambs at 5,7,8 and10-week-old were( 111. 00±16. 97) per lamb,( 139. 50 ± 28. 99) per lamb,( 108. 50 ± 17. 68) per lamb and( 42. 00 ± 11. 31) per lamb,respectively. The number of oocytes obtained oocytes from 5,7 and 8-week-old lambs was significantly higher than that from 10-week-old lambs( P< 0. 05),but there was no significant difference among 5,7 and 8-week-old lambs( P> 0. 05). The maturation rate of oocytes obtained from the test maturation solution was 3. 64 percentage point higher than that from the basic mature solution. The cleavage rate of oocytes from fertilization medium Ⅰ was significantly higher than that from fertilization medium Ⅱ( P< 0. 01). The results indicated that selection of lambs aged 5-8 weeks,oocytes in vitro maturation solution containing a certain amount of cysteine and fertilization medium containing a certain amount of ESS could improve the JIVET system.
引文
[1]KELLY J M,KLEEMANN D O,WALKER S K.Enhanced efficiency in the production of offspring from 4-to 8-week-old lambs[J].Theriogenology,2005,63(7):1876-1890.
[2]RANGEL-SANTOS R,MCDONALD M F,WICKHAM G A.Evaluation of the feasibility of a juvenile MOET scheme in sheep[J].Proc New Zealand Society Anim Proc,1991,51(1):139-142.
[3]KELLY J M,KLEEMANN D O,WALKER S K.The effect of nutrition during pregnancy on the in vitro production of embryos from resulting lambs[J].Theriogenology,2005,63(7):2020-2031.
[4]GOU K M,GUAN H,BAI J H,et al.Field evaluation of juvenile in vitro embryo transfer(JIVET)in sheep[J].Anim Reprod Sci,2009,112(3/4):316-324.
[5]BAI J H,HOU J,GUAN H,et al.Effect of 2-mercaptoethanol and cysteine supplementation during in vitro maturation on the developmental competence of oocytes from hormone-stimulated lambs[J].Theriogenology,2008,70(5):758-764.
[6]刘宗正,李和刚,程明,等.利用JIVET技术快速繁育萨福克羊的研究[J].畜牧与饲料科学,2016,37(3):1-4.
[7]张兴会,张世伟,宋先忱,等.幼龄母羊超数排卵及卵母细胞成熟效果分析[J].中国畜牧兽医,2011,38(11):136-139.
[8]李英,侯健,郑云胜,等.肝素和羊血清对绵羊附睾精子体外获能的影响[J].内蒙古农牧学院学报,1998,19(4):20-24.
[9]马世科,乔海生,马利青,等.幼畜超数排卵及体外胚胎生产技术在青海高原地区的应用研究[J].青海畜牧兽医杂志,2015,45(2):13-14.
[10]陈晓勇,田树军,桑润滋,等.诱导幼羔卵泡发育及体外胚胎生产[J].农业生物技术学报,2008,16(3):456-460.
[11]张锁链,刘东军,王建国,等.通过体外受精技术缩短绒山羊育种周期的研究[J].内蒙古大学学报(自然科学版),1998,29(1):114-117.
[12]李运生,曹鸿国,刘亚,等.羔羊超数排卵与胚胎体外生产技术研究进展[J].动物医学进展,2011,32(11):99-104.
[13]于勇,钟旭,王芬露,等.不同月龄绵羔羊卵泡发育的比较研究[J].中国畜牧杂志,2012,48(9):17-21.
[14]王超,郝泽东.不同培养条件、培养时间和温度对山羊卵母细胞体外成熟的影响[J].西北农业学报,2007,16(2):7-9.
[15]白佳桦.性成熟前绵羊、牛JIVET技术研究[D].北京:中国农业大学,2008.
[16]汪立芹,何宗霖,林嘉鹏,等.萨福克羔羊卵泡诱导发育效果的研究[J].中国农业大学学报,2015,20(4):141-146.
[17]郭洪,万鹏程,石文艳,等.不同品种羔羊超数排卵及胚胎体外生产技术研究[J].畜牧与兽医,2012,44(10):20-24.
[18]吴伟伟,徐新明,张廷虎,等.影响新疆细毛羊羔羊超排及体外受精效果的研究[J].中国畜牧兽医,2011,38(4):144-147.
[2]RANGEL-SANTOS R,MCDONALD M F,WICKHAM G A.Evaluation of the feasibility of a juvenile MOET scheme in sheep[J].Proc New Zealand Society Anim Proc,1991,51(1):139-142.
[3]KELLY J M,KLEEMANN D O,WALKER S K.The effect of nutrition during pregnancy on the in vitro production of embryos from resulting lambs[J].Theriogenology,2005,63(7):2020-2031.
[4]GOU K M,GUAN H,BAI J H,et al.Field evaluation of juvenile in vitro embryo transfer(JIVET)in sheep[J].Anim Reprod Sci,2009,112(3/4):316-324.
[5]BAI J H,HOU J,GUAN H,et al.Effect of 2-mercaptoethanol and cysteine supplementation during in vitro maturation on the developmental competence of oocytes from hormone-stimulated lambs[J].Theriogenology,2008,70(5):758-764.
[6]刘宗正,李和刚,程明,等.利用JIVET技术快速繁育萨福克羊的研究[J].畜牧与饲料科学,2016,37(3):1-4.
[7]张兴会,张世伟,宋先忱,等.幼龄母羊超数排卵及卵母细胞成熟效果分析[J].中国畜牧兽医,2011,38(11):136-139.
[8]李英,侯健,郑云胜,等.肝素和羊血清对绵羊附睾精子体外获能的影响[J].内蒙古农牧学院学报,1998,19(4):20-24.
[9]马世科,乔海生,马利青,等.幼畜超数排卵及体外胚胎生产技术在青海高原地区的应用研究[J].青海畜牧兽医杂志,2015,45(2):13-14.
[10]陈晓勇,田树军,桑润滋,等.诱导幼羔卵泡发育及体外胚胎生产[J].农业生物技术学报,2008,16(3):456-460.
[11]张锁链,刘东军,王建国,等.通过体外受精技术缩短绒山羊育种周期的研究[J].内蒙古大学学报(自然科学版),1998,29(1):114-117.
[12]李运生,曹鸿国,刘亚,等.羔羊超数排卵与胚胎体外生产技术研究进展[J].动物医学进展,2011,32(11):99-104.
[13]于勇,钟旭,王芬露,等.不同月龄绵羔羊卵泡发育的比较研究[J].中国畜牧杂志,2012,48(9):17-21.
[14]王超,郝泽东.不同培养条件、培养时间和温度对山羊卵母细胞体外成熟的影响[J].西北农业学报,2007,16(2):7-9.
[15]白佳桦.性成熟前绵羊、牛JIVET技术研究[D].北京:中国农业大学,2008.
[16]汪立芹,何宗霖,林嘉鹏,等.萨福克羔羊卵泡诱导发育效果的研究[J].中国农业大学学报,2015,20(4):141-146.
[17]郭洪,万鹏程,石文艳,等.不同品种羔羊超数排卵及胚胎体外生产技术研究[J].畜牧与兽医,2012,44(10):20-24.
[18]吴伟伟,徐新明,张廷虎,等.影响新疆细毛羊羔羊超排及体外受精效果的研究[J].中国畜牧兽医,2011,38(4):144-147.
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