【作者】 张兆旺；

【导师】 刘孟洲；

【作者基本信息】 甘肃农业大学 ， 动物遗传育种与繁殖， 2005， 博士

【摘要】 迄今为止,国内外虽有不少有关鸡精液稀释液研究的相关报道,但关于精液稀释液设计与筛选方法学的研究尚属缺乏。在这一领域没有一套完整而实用的理论体系,对诸如精液稀释液pH值、渗透压、缓冲能力和配方物质组成进行调控、筛选和评价,如缓冲能力就没有具体参数可供借鉴,也缺乏测定方法等。有鉴于此,本论文拟从方法学的角度,探索鸡精液稀释液设计与筛选方面的指标体系和测定方法,用以指导制备高受精率的鸡精液稀释液,以适应现代养鸡生产的需要。本论文包括三个部分,第一部分是鸡精液稀释液设计与主要液体质量指标的确定、控制与测定研究。对精液稀释液pH值、渗透压和缓冲能力梯度进行理论控制,然后以实际测定结果检验控制的准确性。第二部分是鸡精液稀释液实验室筛选研究——稀释精液体外精子生存质量与形态学观察研究。通过研究不同pH值、渗透压、缓冲能力和溶质种类的稀释液以及保存时间和温度对保存于其中的精子形态和机能的影响,确定最佳稀释液的合理参数。第三部分是鸡稀释精液人工授精应用研究,以稀释精液进行人工授精,以受精率对稀释液及其设计和筛选方法的优劣做出客观的评价。实验结果表明:①pH值的理论控制准确率在98.69%,渗透压准确率在95.9%-99.1%,缓冲指数准确率在97.6%以上。②在2℃-5℃低温条件下,稀释液缓冲指数不影响精子活率,但影响精子畸形率。缓冲指数348时,精子畸形率极显著地低于缓冲指数119.6～228.8和461.6～1147.6范围的畸形率。为了保持精子正常的形态,不但要求要有合适的pH值和渗透压,同时还要求有适中的缓冲指数。稀释液缓冲指数对精子畸形率的影响较精子活率的影响大。③柠檬酸钠、葡萄糖对鸡精液的保存,具有重要作用,而氯化钾在鸡精液稀释液中并不是必需的;复方稀释液保存效果普遍优于单方稀释液;低温下鸡精液稀释液渗透压应为等渗或接近等渗,且最适渗透压还与构成稀释液的物质种类和相互配比有关系;稀释液PH值应接近中性,最好与精液pH值保持一致;在缓冲指数为3.894-7.257的范围内,缓冲指数与精子保存效果的两个指标均呈现显著的负相关关系。④精子最高生存指数出现在三种有机盐的稀释液中;两种无机盐液中精子存活时间最长,但精子畸形率很高;渗透压值与精子头部畸形类型高度相关(p<0.01),与精子头部弯曲且膨胀百分数的相关系数为-0.93,与精子头部单纯弯曲百分数的相关系数为0.94。在300mmol/L时,精子生存指数、存活时间均较好;500mmol/L时精子畸形率最低;渗透压对保存效果有极显著的影响,物质种类对保存48h精子形态有显著地影响,且渗透压与精子畸形类型有一更多还原

【Abstract】 So far, there are many studies on chicken semen extender, but the study on the methodology of design and riddling of it is a gap. There is not integrated and applied system on this field, so that the pH,osmotic pressure (OP), buffer capacity (BC) and material of semen extender were controlled , riddled and valued, for example, there is not idiographic parameter on the BC used for reference and there is not measure method, too. So the index system and measure method of chicken semen extender design and riddling were researched in present study on the methodology point of view, in order to confect semen extender that lead to high fertility and adapt the need of modern chicken production.The present study was divided to three parts. The first part included designing of chick semen extender and ascertaining, controlling and measuring the solution quality. The pH, OP and buffer capacity grads (BCA) of semen extender were controlled theoretically, and then exam the nicety rate according the result measured actually. The second part of the study was determining the composition of extender in lab by sperm quality and morphology in it in vitro. The sperm was stored in extenders which their pH,OP,BCA were different for varied time and temperature, and observed the effect on sperm, in order to determine the optimal parameter of the optimized semen extender. The third part was the research of application of AI using diluted semen. According to AI in practically production and the fertility rate of eggs,the extenders were valued objectively.The results showed that:①The nicety rate of the pH was 98.69% and the osmotic pressure was 95.99%–99.1%, the grads of buffer index of extenders were more than 97.6%.②The buffer index (BI) did not affect the motility of sperm, but did the abnormality ratio. The abnormality ratio was lower significantly when the buffer index was 348 than 119.6～228.8 and 461.6～1147.6.In order to keep the normal shape of sperm, the pH and osmotic pressure should be suitable, at the same time, the buffer index also be moderate. The effect of buffer index on abnormality ratio was greater than sperm motility.③First, sodium citrate, glucose play an important role in chick semen extender, but the Potassium chloride (PCH) is not necessary. Second, compound extender is better than the simple one. Third, at low temperature, the osmotic pressure should be equal the osmotic pressure of cock semen or be close to it, what is more, the optimum osmotic pressure is related with the material that compose extender and their proportion in it. Fourth, the pH should be litmusless or be same with semen. Fifth, when buffer index range from 3.894 to 7.257, there are significant negative correlation between buffer index and the two index of semen storage.④The tiptop survival index of sperm showed in three kinds of organic salt extenders; the longest survival time appears in two inorganic extenders, but the monstrous sperm is more; in the extender containing sucrose and monosodium glutamate (MG), the abnormality sperm only 5.29% and 5.40%; there are high correlation between osmotic pressure and monstrous sperm head (p<0.01), the correlation coefficient was -0.93 with flexural and bulgy sperm head , and 0.94 with only flexural one. When the concentration was 300mmol/L, sperm survival index (SSI) and time (SST) were better; and monstrous sperm was the lowest when 500mmol/L. The effect of osmotic pressure on更多还原