【作者】 魏世宝；

【导师】 金一；

【作者基本信息】 延边大学 ， 动物遗传育种与繁殖， 2007， 硕士

【摘要】 本次研究的主要目的是评估冷冻稀释液中添加不同的抗氧化剂对冻精解冻后精子质量参数、膜脂质过氧化程度及体外受精率的影响。以手握法采集的猪精液为试验材料，进行处理后，用不同规格的细管进行了冷冻试验。试验主要分3步进行：Ⅰ将精液按不同的比例稀释，以不同的速度离心后，除去上清液，再用冷冻稀释液进行稀释，然后用不同容量的细管进行冷冻，解冻后对精液品质进行生物学检测；Ⅱ在冷冻稀释液中添加N-乙酰半胱氨酸、L-半胱氨酸和N-乙酰半胱氨酸+L-半胱氨酸混合物，配制成不同的冷冻稀释液，冷冻解冻后对精液品质进行生物学检测。Ⅲ在冷冻稀释液中添加N-乙酰半胱氨酸、L-半胱氨酸和N-乙酰半胱氨酸+L-半胱氨酸混合物，用细管进行冷冻解冻后，对膜脂质过氧化反应的程度进行生物学检测。试验结果如下：1、800转／分离心后，精子的活率、顶体完整率显著高于500转／分和1000转／分的，而低渗膨胀率却显著低于500转／分和1000转／分的(P＜0.05)；1000转／分离心后，精子的顶体完整率显著降低，而低渗膨胀率却大大地提高(P＜0.05)。虽然，不同的离心速度对运动学参数没有影响(P＞0.05)，但是800转／分的离心效果要好于500转／分和1000转／分的。2、精液1：1稀释与1：2稀释的结果，其冷冻后精子的活率差异不显著(P＞0.05)；但1：1稀释的其冷冻后精子的顶体完整率显著高于1：2稀释的(超过了15个百分点)，而低渗膨胀率却显著低于1：2的(10个百分点)；在运动学参数方面，只有LIN显著降低(P＜0.05)，而对其它的运动学参数没有影响(P＞0.05)。从冷冻所使用的细管来看，两种不同比例稀释后，在其各方面的指标中，0.5ml细管的冷冻效果要优于0.25ml细管的，但两者之间差异不显著(P＞0.05)。3、与对照组相比，在冷冻稀释液中添加1mM的N-乙酰半胱氨酸能显著地提高解冻后精子的活率和顶体完整率，降低精子的低渗膨胀率(P＜0.05)。在运动学参数方面，只有VAP显著升高(P＜0.05)，而对其它的运动学参数没有影响(P＞0.05)。体外胚胎生产的数据显示：在冷冻稀释液中添加不同浓度的N-乙酰半胱氨酸对卵裂率没有影响(P＞0.05)。4、与对照组相比，在冷冻稀释液中添加0.1mg／ml的L-半胱氨酸能显著地提高解冻后精子的活率和顶体完整率，降低精子的低渗膨胀率(P＜0.05)，而对运动学参数没有影响(P＞0.05)。体外胚胎生产的数据显示，与对照组、0.2mg／ml处理组和0.3mg／ml处理组相比，在冷冻稀释液中添加0.1mg／ml的L-半胱氨酸可以显著地提高卵裂率(P＜0.05)。5、与对照组相比，在冷冻稀释液中添加1mM N-乙酰半胱氨酸+0.1mg／ml L-半胱氨酸能显著地提高解冻后精子的活率和顶体完整率，降低精子的低渗膨胀率(P＜0.05)。在运动学参数方面，只有LIN降低，而VAP、ALH和BCF都显著提高(P＜0.05)。体外胚胎生产的数据显示：与对照组相比，在冷冻稀释液中添加1mM N-乙酰半胱氨酸+0.1mg／ml L-半胱氨酸，可以显著地提高卵裂率(P＜0.05)。6、与对照组相比，在冷冻稀释液中添加不同浓度的L-半胱氨酸，膜脂质过氧化水平明显降低(P＜0.05)；而添加N-乙酰半胱氨酸和L-半胱氨酸+N-乙酰半胱氨酸，对膜脂质过氧化反应没有影响(P＞0.05)。更多还原

【Abstract】 The aim of this study is to examine the effect of addition of different antidants onquality parameters of frozen-thawed boar semen, membrane lipid peroxidation and theability of in vitro embryo production. With holding the penis to obtain pig semen asmaterial, this text carried on the experment of the boar frozen, semen that was frozen bythe straw. The research contents have three respects: I The semen was diluted indifferent proportion, removed clear liquid after being concentrated at different speeds,and then diluted the semen with freezing extender, and finally frozert with the differentcapacity straws, measured the quality of semen by biological method after postthaw;ⅡThe frozen semen that was respectively made with the different kinds of extenders,adding NAC, L-cysteine, and NAC plus L-cystein, measured the quality of semen bybiological method after postthaw;ⅢEffects of the additions of NAC, L-cysteine andNAC plus L-cysteine in the freezing extender on the membrane lipid peroxidation weremeasured by biological method. All resnlts of the test are as follows:1、The motility and acrosomal integrity of 800r/m centrifugation is significant higherthan 500 r/m and 1000r/m, while the tail-coiling is significant lower than 500 r/m and1000r/m (P＜0.05). 1000r/m centrifugation, the acrosomal integrity is significant lowerthan 500 r/m and 800r/m, while the tail-coiling is significant higher than 500 r/m and800r/m (P＜0.05). Although different centrifuge can not affect sperm kinematicparameters (P＞0.05), but the results of 800r/m centrifuge is significant better than500r/m and 1000r/m.2、The differences of the thawed sperm motility are not significant with 1:1 and 1:2diluting semen before centrifuge. (P＞0.05). But the acrosomal integrity 1:1 dilutingsemen is significantly higher than that of 1:2 one after freezing (exceed 15 percentagepoints); tail-coiling is lower extremely (percentage points) of 1:1 diluting semen than1:2 one after freezing (P＜0.05). In.relation to the sperm kinematic parameters, only LIN was affected by the different proportion(P＜0.05). From using different straws, after twodifferent proportions dilute, 0.5ml is superior to 0.25ml among of various field indexes,but the difference is not remarkable between the two (P＞0.05).3、The addition of 1mM NAC in sperm extender for freezing significantly increased thepost-thaw rates of sperm motility and acrosomal integrity compared with the controlgroup, decreased the postthaw rate of sperm tail-coiling (P＜0.05). In relation to spermmotion kinetics, only VAP was affected by the presence of NAC in the freezing extender,being significantly higher in spermatozoa frozen in the presence of NAC(P＜0.05). Datafrom in vitro embryo production showed that the addition of NAC in the freezingextender had no significant effect on the rate of cleaved zygotes (P＞0.05).4、The addition of 0.1mg/ml L-cysteine in sperm extender for freezing significantlyincreased the postthaw rates of sperm motility and acrosomal integrity compared withthe control group, decreased the postthaw rate of sperm tail-coiling (P＜0.05). However,L-cysteine did not affect any of the sperm kinematic parameters evaluated (P＞0.05).Data from in vitro embryo production revealed an improvement of cleavage rates whenspermatozoa were frozen in the presence of 0.1mg/ml L-cysteine compared with controlor the 0.2 mg/ml L-cysteine and 0.3 mg/ml L-cysteine groups (P＜0.05).5、The combined addition of NAC and L-cystein in the sperm extender for freezingsignificantly increased the postthaw rates of sperm motility and acrosomal integritycompared with the control group, decreased the postthaw rate of sperm tail-coiling(P＜0.05). In relation to the sperm kinematic parameters, VAP, ALH, and BCF werehigher and LIN lower in thaw spermatozoa frozen in the presence of NAC plusL-cystein compared with controls (P＜0.05). Data from in vitro embryo productionshowed, when compared with controls, a significantly higher proportion of zygotes(P＜0.05).6、The presence of L-cystein in the freezing extender significantly decreased the level ofmembrane lipid peroxidation (P＜0.05), while the membrane lipid peroxidation wasunaffected by the presence of NAC, NAC plus L-cystein in the freezing extender(P＞0.05).更多还原