不同染色与冷冻方法对关中奶山羊X、Y精子的分离及应用效果研究

为探究不同Hoechst 33342的染色浓度与冷冻方法对关中奶山羊X、Y精子进行分离及应用效果的影响，本实验采集6只关中奶山羊精液，使用浓度分别为10、11、12、13μL/m L的Hoechst 33342(5 mg/m L)染色液对精液样本进行染色，以流式细胞仪分离X精子和Y精子，应用3种不同冷冻程序冷冻精液，应用体外受精与胚胎发育技术评估关中奶山羊X、Y精液的分离准确性。结果表明：12μL/m L Hoechst 33342染色条件下，奶山羊X、Y精子分离效率达到91.02%，高于10μL/m L和13μL/m L(P<0.01)，最高分选速度为4 134个/s(P<0.05)；公羊个体的精液品质越好，其精子的分离速度越快，但对分离准确率的差异不显著；使用冷冻程序2进行关中奶山羊的X、Y精子冷冻的效果较好（P<0.05）；体外受精48 h后，使用Y精子进行受精的卵细胞的卵裂率高于使用X精子（P<0.05），在胚胎培养第9天，使用Y精子进行受精的卵细胞的囊胚发育率高于使用X精子（P<0.05）。总之，当Hoechst 33342浓度为12μL/m L...     

广州地区娟姗公牛与荷斯坦公牛精液品质的比较研究

本试验选择亚热带气候条件下广州地区的娟姗公牛和荷斯坦公牛各5头,比较两个品种公牛的精液品质(采精量、原精密度、原精活力、细管精液产量、冻后活力、低渗膨胀率及穿透率)。研究表明,荷斯坦公牛每次采精的采精量(16.14±0.06 mL)和细管精液产量(189.17±3.11支)都极显著地高于娟姗公牛(4.74±0.05 mL,158.46±2.64支)(P<0.01);娟姗公牛的原精密度(8.95±0.08亿/mL)极显著地高于荷斯坦公牛(8.32±0.07亿/mL;P<0.01);娟姗公牛原精活力(0.731±0.004)高于荷斯坦公牛(0.729±0.003),但两者差异不显著(P<0.05);娟姗公牛精液的冻后活力(0.355±0.003)极显著高于荷斯坦公牛(0.339±0.003;P<0.01);娟姗公牛冷冻精液的低渗膨胀率(34.50%±0.49%)显著高于荷斯坦公牛(31.21%±0.59%;P<0.01);娟姗公牛冷冻精液对去透明带仓鼠卵的穿透率(84.51%±13.83%)显著高于荷斯坦公牛(81.52%±6.13%;P<0.05)。     

梅花鹿XY精子分选及冷冻精液制备

为分选梅花鹿XY精子及制备冷冻精液,2006年9月21~29日,对电刺激采集的梅花鹿精液经过长距离运输,流式细胞仪分离,冷冻,解冻后精液重分析和品质鉴定。结果显示:常规冻精和X、Y型冻精性别比分别为50%、91.21%和94.08%,常规冻精与X、Y型冻精组间差异显著(P<0.05);活率分别为0.43±0.04、0.45±0.04和0.43±0.03,常规冻精与X、Y型冻精组间差异不显著(P>0.05);存活指数分别为0.25、0.05和0.05,常规冻精与X、Y型冻精组间差异显著(P<0.05);顶体完整率分别为(77.6±1.0)%(、84.6±0.5)%和(82.2±0.6)%常规冻精与X、Y型冻精组间差异不显著(P>0.05)。用流式细胞技术可以分选获得梅花鹿XY冷冻精液,性别比和品质达到低剂量人工授精的要求。     

荷斯坦公牛X和Y精子核形态差异及其影响因素分析

旨在分析荷斯坦公牛X和Y精子核形态(包括精子大小和形状)及其差异,探究流式分选及分选后冷冻对公牛X和Y精子核形态的影响.本研究使用荧光显微镜拍摄15头年龄相当、健康状况良好的荷斯坦公牛X和Y精子冷冻前后及新鲜精液分选前后共6类精子核图像,利用Image J插件Nuclear Morphology Analysis分析3...     

应用两种稀释液冷冻驴精液试验报告

本试验选用两种冷冻稀释液,通过离心去除精清的方法对驴精液进行了冷冻。结果发现在试验条件相同的情况下,用配方2冷冻稀释液冷冻精液的效果好于配方1,其平衡后活率(0.700±0.038)极显著高于配方1的平衡后活率(0.605±0.040)(P<0.01);经过冷冻后,其冻后活率(0.460±0.066)也极显著高于配方1的冻后活率(0.325±0.064)(P<0.01);且其精子顶体完整率(0.800±0.031)也极显著高于配方1(0.736±0.024)(P<0.01)。这表明配方2是1个冷冻驴精液的较好配方。     

绒山羊精液冷冻保存技术的研究

为了使绒山羊精液得到长期保存,充分发挥和提高优良种公羊的利用率,本实验对绒山羊精液冷冻保存技术进行了研究,比较了4种自行研制的保存稀释液的冷冻效果。结果表明:采用Ⅰ液(葡萄糖-卵黄-柠檬酸钠)冷冻保存绒山羊精液,解冻后精子活率(0.508±0.010)极显著高于Ⅱ液(葡萄糖-卵黄-乳糖-柠檬酸钠)(0.275±0.008)、Ⅲ液(葡萄糖-卵黄-蔗糖-柠檬酸钠)(0.354±0.012)和IV液(葡萄糖-卵黄-蔗糖-乳糖-柠檬酸钠)(0.319±0.006)(P<0.01);Ⅲ液极显著高于Ⅱ液(P<0.01)和显著高于Ⅳ液(P<0.05)。解冻后Ⅰ液的精子顶体完整率(59.4%±0.5%)极显著高于Ⅱ液(47.1%±0.8%)、Ⅲ液(52.4%±0.6%)和IV液(51.3%±0.5%)(P<0.01)。精液解冻后在室温(23±2)℃下避光培养,Ⅰ液中精子活率在5h内能够保持0.30以上,Ⅱ、Ⅲ液和Ⅳ液精子活率维持在0.3不到2h。     

物候节律对河流型水牛精液质量影响的分析

对摩拉水牛和尼里水牛2006—2014年不同季节和节气的射精量、原精活力、精子密度、解冻活力进行测定、记录和统计分析。结果:河流型水牛春季的射精量、精子密度、原精活力和冷冻精液解冻活力平均分别为(4.80±2.063)m L(n=2 216)、(57.61±14.727)%(n=2 209)、(7.79±4.165)亿/m L(n=2 213)和(36.00±2.581)%(n=1 413);夏季的射精量、精子密度、原精活力和冷冻精液解冻活力平均分别为(53.28±2.311)m L(n=2 392)、(60.32±10.515)%(n=2 379)、(8.26±4.250)亿/m L(n=2 389)和(35.99±2.506)%(n=1 823);秋季的射精量、精子密度、原精活力和冷冻精液解冻活力平均分别为(5.28±2.161)m L(n=1 977)、(61.15±16.462)%(n=1 965)、(8.28±4.212)亿/m L(n=1 977)和(36.56±9.897)%(n=1 517);冬季的射精量、精子密度、原精活力和冷冻精液解冻活力平均分别为(5.02±2.019)m L(n=2 350)、(57.11±18.910)%(n=2 337)、(7.68±3.901)亿/m L(n=2 344)和(36.21±2.615)%(n=1 476)。     

比利时蓝白花牛XY精子分选及冷冻精液制备

本研究利用流式细胞仪分选比利时蓝白花牛X、Y精子并制备冷冻精液。选择平均年龄4岁的健康比利时蓝白花种公牛12头，假阴道法采集精液送至实验室分选，经流式细胞仪分离、冻存、解冻后精液重分析和品质鉴定。X、Y冻精（93.4%和91.1%）性别比例显著高于常规冻精（50%）；分选后精液冻后存活率、活力和顶体完整率与常规冻精差异不显著。本研究结果显示，分选前控制公牛的精液品质（活力≥70%，畸形率≤18%）可以明显改善分选效果；分选后精子纯度和冻后活力满足低剂量人工授精要求（纯度>90%，活力>30%），精子分选对比利时蓝白花牛产业的发展具有重要意义。     

夏季高温对不同品种种公牛精液生产指标的影响

选择荷斯坦、夏洛来、利木赞、西门塔尔、皮埃蒙特种公牛各2头作为实验牛,研究夏季高温对各品种种公牛精液生产指标的影响差异。结果表明:①从采精量来看,夏洛来的每次采精量极显著(P<0.01)高于西门塔尔、利木赞和皮埃蒙特,显著(P<0.05)高于荷斯坦。②从原精液的密度来看,利木赞的原精液密度极显著(P<0.01)高于夏洛来、西门塔尔、皮埃蒙特。荷斯坦的原精液密度极显著(P<0.01)高于夏洛来、西门塔尔、皮埃蒙特。③从冻精生产数来看,荷斯坦与夏洛来、西门塔尔、利木赞、皮埃蒙特存在极显著(P<0.01)差异。夏洛来与皮埃蒙特差异极显著(P<0.01)。     

冻融过程对绵羊精子ATP酶活性的影响

为了充分发挥和提高优良种公羊的利用率,提高绵羊冻融精液品质,试验采用酶动力学分光光度法对冷冻保存前后小尾寒羊精子和精清中ATP酶活性变化进行研究。结果表明:冷冻保存小尾寒羊精子中ATP酶活性(83.76±3.53)U/m L与冷冻前(175.42±4.32)U/m L相比极显著降低(P0.01),精清中ATP酶活性(164.96±3.22)U/m L与冷冻前(73.51±2.23)U/m L相比极显著升高(P0.01);冷冻前后精子活率与精子ATP酶活性均呈极显著正相关(r=0.979和0.968,P0.01)。说明冻融过程对绵羊精子中ATP酶的活性造成了严重损伤。绵羊冻融精子ATP酶的活性可以在一定程度上预测精子活率,并可作为绵羊精液品质评价的重要指标。     

性别控制与体外受精技术快速繁育高产奶牛群

使用流式细胞分离仪可以将X精子从Y精子中选离出来,分离精度高达90%或更高。这种性别控制(性控)技术可以用来生产性别预选的家畜以及优化育种的方案。然而,精子分离的速度以及其性控冷冻精液的不同受精力仍然限制着该技术的广泛应用,尤其是在传统的人工授精育种方案中。体外受精技术(Invitro fertilization,IVF)提供了一个可行而有效的手段来提高奶牛性控精子的受精效率。本文介绍了有关利用IVF技术来控制奶牛性控胚胎发育率和提高雌性胚胎质量的新方法,从而提高胚胎生产效率,达到快速繁育高产奶牛群的目的。这些IVF程序包括以优化的方法遴选出用每头公牛X精子进行IVF所需的最佳肝素浓度;减少IVF受精液的体积以确定每枚卵子获得受精所需的最少精子数目;在体外培养液中使用果糖来替代葡萄糖,以及使用能超速降温的玻璃化冷冻技术。     

低剂量梅花鹿XY精子马花杂交人工授精试验

采用流式细胞分离仪分离的梅花鹿X和Y型冷冻精液与常规冻精对62头3组同期化处理的马鹿进行直肠把握人工授精。结果表明,0.25 mL/支含106个有效精子的X和Y型冻精产仔率分别为43%和37%,而0.25 mL/支含107个有效精子的常规冻精产仔率为55%,X、Y型冻精与常规冻精组间差异显著(P<0.05),所产后代性别比率分别为0∶10,9∶0和5∶6,X、Y型冻精与常规冻精组间差异显著(P<0.05),X、Y型冻精与常规冻精所产后代出生及60 d时的体重差异不显著(P>0.05)。     

Influence of Random Factors and Two Different Staining Procedures on Computer-assisted Sperm Head Morphometry in Bulls

Contents Computer-assisted sperm analysis has the potential to improve reproducibility and objectivity in the assessment of sperm morphology. The aim of this study was to evaluate the use of a computer-assisted sperm morphometry analysis system for the determination of sperm head dimensions in bulls. Two experiments were performed to determine the variability caused by random factors and the influence of two different staining procedures. In the first experiment, three ejaculates were collected from each of five clinically healthy bulls. Air-dried semen smears were stained using a modified Farelly staining. The slides were observed via bright field microscopy with green filter using a 100× oil immersion objective. A video camera attached to the microscope transmitted images to a personal computer. Each sperm head was identified and analysed by the computer software (Morphology Analyzer V. 1.5; Mika Medical GmbH, Ismaning, Germany). Area, length and width of each sperm head were calculated and stored in a database for further statistical analysis. A minimum of 100 sperm heads were evaluated per slide. In experiment 2, the influence of two different staining procedures (Farelly and Papanicolaou) on sperm head dimensions was determined. The mean spermatozoal head measurements across all slides for area, length and width were 40.49 μm², 9.70 μm and 5.30 μm, respectively. On the basis of these results, the variability between slides, ejaculates and bulls using variance component estimation was calculated. All random factors (bull, ejaculate and slide) had a significant effect (p < 0.001) on sperm head dimensions. However, the variability attributable to bull (18.89–51.72%) was considerably higher compared with that of slide and ejaculate (0.17–5.27%). Additionally, differences existed between bulls concerning the shape and normality of histograms of their sperm head dimensions. The minimum number of spermatozoa required for analysis of sperm head dimensions was found to be about 60 spermatozoa per sample. The use of Papanicolaou stain resulted in significantly smaller sperm head dimensions, e.g. sperm head area 31.48 versus 38.35 μm² (p < 0.001). In conclusion, computer-assisted sperm head morphometry provides an objective, precise and reproducible tool. Comparisons of results from different studies should consider the influence of random and experimental factors to avoid misinterpretation.     

The Effects of Cryopreservation on the Morphometric Dimensions of Iberian Red Deer (Cervus elaphus hispanicus) Epididymal Sperm Heads

Computer-automated sperm-head morphometry was used in this study to determine the effects of cryopreservation on red deer sperm-head morphometry. Epididymal sperm samples were collected from 40 mature stags and were divided. One portion was diluted at room temperature in a Tris-citrate egg yolk medium, containing 6% glycerol. A microscope slide was prepared from single extended sperm samples prior to freezing. The remainder of each sample was frozen in nitrogen vapours. After thawing, sperm smears were prepared as described above. All slides were air dried and stained with Hemacolor. The sperm-head dimensions for length, width, area, perimeter and shape factor (length/width), for a minimum of 135 spermatozoa were determined for each slide by means of the Sperm-Class Analyser (SCA). Firstly, our results show that cryopreservation substantially reduced (p < 0.001) sperm motility and plasma membrane and acrosome integrities. In addition, sperm heads were significantly smaller in cryopreserved spermatozoa than in the companion extended samples for area (32.05 microm2 vs 32.56 microm2; p < 0.05), length (8.46 microm vs 8.53 microm; p < 0.0001) and shape factor (1.833 vs 1.849; p < 0.0001) for all stags. These differences were found within 29 of 40 stags (75%) for at least three of the morphometric parameters. The individual variability (CV) of sperm head measurements from extended samples was negatively correlated (p < 0.005) with the per cent of change in sperm head measurements after cryopreservation for area (r = -0.465), width (r = -0.483) and perimeter (r = -0.375). Thus, the lower the sperm head variability in the extended samples, the greater the sperm change as a consequence of the cryopreservation. These results suggest that the variability (heterogeneity) in sperm head dimensions of individual stags may be a good indicator of sperm freezability.     

茸鹿人工选育品种品系数量性状遗传参数的统计分析

对我国人工选育的梅花鹿和马鹿 6个品种品系的 2 0多个数量性状的遗传参数 ,进行了统计分析 ,结果表明 ,变异系数除鲜茸重的较高之外 ,其他很小 ;公鹿的留种率很低 ,茸重性状的选择差很大 ;遗传力和重复力均较高 ;世代间隔较短 ;鲜茸重的遗传进展和年改进量较大 ;梅花鹿种公鹿的种用年限较短 ,并明显低于马鹿的 ;梅花鹿品种品系间和马鹿亚种间杂交F1茸重性状和繁殖成活率性状的杂种优势率非常显著 (P <0 0 1 )。此项统计分析为茸鹿的优质高效育种、提高纯繁选育速度、杂种优势利用、杂交培育新品种和育种规化及模式提供了科学依据。     

Associations between sperm abnormalities, breed, age, and scrotal circumference in beef bulls

The objectives of this study were to determine the associations of breed, age, and scrotal circumference (SC), and their interaction, on the prevalence of sperm abnormalities in beef bulls in Alberta, Canada, and the percentage of satisfactory potential breeders identified during breeding soundness examination solely due to normal sperm morphology. Eosin-nigrosin stained semen smears and evaluation reports of 1642 bull breeding soundness evaluations were procured from 6 veterinary clinics in Alberta. Sperm morphology was determined for at least 100 sperm per bull. The most common defects were detached head [4.86% ± 5.71%; mean ± standard deviation (s)], distal midpiece reflex (6.19% ± 9.13%), and bent tail (1.01% ± 1.54%). Although breed, age, and SC did not significantly affect the prevalence of head or midpiece defects, morphologically normal or abnormal sperm, tail defects were more prevalent in Angus and Hereford bulls compared with other breeds. Overall, solely on the basis of sperm morphology, 1363 (83.0%) bulls were classified as satisfactory potential breeders and the remainder 279 (17.0%) as unsatisfactory (> 30% abnormal sperm, > 20% defective heads, or both). Although not significantly different, the breed with the highest percentage of satisfactory potential breeders was Limousin (90.6%) and the lowest was Hereford (78.8%). That 17% of bulls subjected to breeding soundness evaluation were designated as unsatisfactory solely on the basis of sperm morphology highlights its importance.     

马鹿精子形态与超微结构研究

文中对马鹿(Cervus elaphus)精子形态与超微结构进行了研究。马鹿精子经固定、脱水、置换、包埋和聚合,用超薄切片机切片,再用醋酸双氧铀、柠檬酸铅染片,最后于透射电镜下观察其超微结构的变化。观察结果表明:马鹿精子全长(58.75±2.35)μm,由头部(8.93±0.24)μm、颈部(1.00±0.16)μm和尾部(48.18±1.18)μm三部分组成;头部呈扁卵圆状,绝大部分被浓缩且电子密度较高的精核所占据,顶体似帽状扣在精核之上,约占头部的2/3,其前部较为膨大;颈部位于头部和尾部之间,这个区域较短;尾部可分为中段、主段和终段。马鹿尾部轴丝的结构类型为"9+2";微管结构类型为"9+9+2"。     

分离Y精子在茸鹿性别控制中的应用研究

为研究一种新的茸鹿繁殖方法,2007～2008年对469头马鹿,79头梅花鹿进行性别控制技术研究。结果表明：性别控制冻精情期受胎率马鹿为87．21％,梅花鹿为60．76％,两者之间差异极显著（P〈0．01）,性比率（指产公仔率）马鹿为90．95％,梅花鹿为93．75％,两者之间无显著差异（P〉0．05）;性别控制冻精与普通冻精输精比较,马鹿情期受胎率之间、性比率之间和梅花鹿性比率之间都存在极显著差异（P〈0．01）,梅花鹿情期受胎率之间则无显著差异（P〉0．05）。普通冻精受胎率马鹿为94．89％、梅花鹿为61．63％,性比率马鹿为50．22％、梅花鹿为49．06％,性别控制冻精受胎率整体接近普通冻精,性比率则提高40％以上,比较分析认为对茸鹿X、Y精子分离进行性别控制试验是成功的。     

使用流式细胞仪分离精子进行仔猪性别控制的研究

本研究旨在探索流式细胞仪分离精子在猪性别控制中的应用。使用流式细胞仪分离猪XY精子,而后通过母猪输卵管授精生产"预知"性别的仔猪,并使用吖啶橙染色法检测粗分离对精子核酸含量的影响。结果,成功利用分离获得的猪X和Y精子对母猪进行输卵管授精,母猪怀孕率、产仔率均为100%;输Y精子母猪产仔雄性率100%(♂6/6),对照母猪产仔雄性率57.14%(♂8/14);3头输X精子母猪产母仔率91.67%(♀11/12),对照母猪产雌性仔猪40%(♀2/5);使用性别分离精子不影响母猪的怀孕率、产仔率,但窝产仔数较低;吖啶橙染色法检测结果表明,流式细胞仪粗分离对猪精子核酸含量没有显著影响(P>0.05)。本研究结果提示,使用流式细胞仪分离精子授精可以有效改变仔猪的性别比例。本研究结果为猪分离精子性别控制技术的推广应用奠定了基础。