低温和超低温保存对中国大鲵成熟精子的影响

中国大鲵Andrias davidianus是中国特有的一种濒危有尾两栖类动物,为了保护这一珍稀物种并且为中国大鲵人工辅助繁育建立一套可靠的技术,对中国大鲵的精子在0—4℃下低温短期保存和在液氮中超低温长期保存进行了研究。研究表明中国大鲵精子原液在常温下一般仅存活3—5h,4℃下能存活6d,0℃下存活时间可达9d。精子原液中添加Ringer氏液或Holtferter氏液,精子存活率比原液保存显著降低。在改良的超低温冻存条件下,中国大鲵精子在液氮中保存两周后解冻,精子复苏率可达10%—15%。利用扫描和透射电镜观察冻融前后中国大鲵精子的超微结构,发现受冷冻损伤精子质膜出现膨胀、破裂;精子穿孔器和轴丝显著弯曲,甚至断裂;波动膜明显脱落或膜结构损坏;精子线粒体嵴变形。结果表明,超低温冻存导致部分中国大鲵精子超微结构发生变化而产生冻伤,进而导致精子活力、复苏率下降。研究为建立规范化的中国大鲵精液保存程序和中国大鲵规模化人工繁育提供重要参考。     

CRYOPRESERVATION OF SEA URCHIN EMBRYOS AND SPERM

A simple method for preserving live sea urchin embryos and sperm in liquid nitrogen (LN) wasdeveloped through the use of DMSO as a cryoprotective additive. Samples of late embryos in double test tubes were cooled to— I96°C by two-step freezing, first to — 76°C and then by plunging in LN. In the case of fertilized eggs, samples were previously frozen to — 40°C, at which temperature the samples were kept for 15 min; they were then transferred into LN. After preservation in LN for various lengths of time, samples in the double test tubes were thawed in water at 15°C. The post-thaw survival was more than 90% for late embryos, and about 10% for fertilized eggs. Difference in the length of the cryopreservation period did not affect survival. Post-thaw development in cryopreserved embryos often showed abnormalities in structure. Sperm with 1.5 M DMSO was successfully preserved in LN by a similar method to the one used for cryopreservation of late embryos. Fertilizability in cryopreserved sperm was complete, regardless of the length of the preservation period. Nearly all the eggs fertilized by cryopreserved sperm developed to normal plutei.     

兰州鲇精液超低温冷冻保存技术研究及细胞损伤检测

为保护兰州鲇(Silurus lanzhouensis Chen)种质资源,促进新品种选育,文章开展了兰州鲇精液超低温冻存抗冻剂筛选及程序保存技术研究,并通过精子解冻激活率、核DNA检测、扫描电镜和透射电镜检测技术对冻存效果和冻存损伤情况进行了评测。结果表明:兰州鲇精液以10%DMSO为抗冻剂, 4℃平衡20min,–80℃平衡30min后立即置于液氮超低温保存,并于40℃水浴解冻时精子冻存效果较佳,冻精激活率达(75.56±3.91)%。SCGE检测DNA损伤结果显示超低温冻存10d、20d及30d兰州鲇精液的彗星率和损伤系数无显著差异。扫描电镜检测显示兰州鲇精子明显分头部,中段及尾部3部分,属于单鞭毛型,无顶体,无侧鳍,中心粒相互垂直呈"T"形,鞭毛为典型的"9+2"微管结构。冻存结构损伤主要表现为膜损伤,细胞质膜与染色质膜发生破损、折皱、囊泡化或整体脱落,细胞核膜与质膜空隙加大,核发生变形,核质疏散,线粒体结构弥散,线粒体内容物外流,中心粒复合体易位,鞭毛外膜变形脱离,中段位置断裂,微管结构基本完好。研究筛选的超低温冷冻保存技术可长期有效保存兰州鲇精液,为兰州鲇种质资源保护及今后...     

猕猴桃简易降温和低乙烯贮藏技术及应用

通过对猕猴桃果实成熟主要指标的测定,确定了秦美适宜采收期为１０月３日至１２日,西选１号和海沃德为１０月下旬。这时果实可溶性固形物含量为６．５─９％,硬度１２─１６ｋｇ／ｃｍ２。猕猴桃贮鲜技术在１９９２／９３年度中试（示范）成功的基础上,１９９３／９４年度在６４ｔ果实上的应用又取得了好结果。秦美贮藏１５７天,果实新鲜,完好果和商品果分别为９６．６％和９７．４％。果实在贮藏期间硬度逐渐降低,由采收时１２─１６ｋｇ／ｃｍ２下降到１．６ｋｇ／ｍ２;淀粉在酶的作用下不断转化为可溶性糖,可溶性固形物由采收时６．５─９％增加到１３．７％。简易降温库成本低,可利用自然低温设施,果实在箱袋内加保鲜剂后置该库内有利于延长猕猴桃贮藏保鲜期。     

中国花鲈精子的超低温冷冻保存及酶活性检测

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疫霉属菌种的液氮超低温保藏

对疫霉属的15个种68株菌和霜疫霉2株菌进行了液氮超低温的保藏试验并得到成功。比较了冷冻速度、保护剂和解冻速度对菌存活的影响。严格控制每分钟降温摄氏1度直到-40℃后再放入液氮罐中,对疫霉和霜疫霉来说都是非常必要的。这种降温程序可通过简单设备人工操作达到。而直接由室温降到-150℃以下会损伤菌种以致死亡。在所用的保护剂中,不论10％甘油还是5-15％二甲基亚砜都能起到保护作用。尚看不出对那种保护剂有特别的要求,似可任意选用。至于解冻条件,由液氮中取出放置在38℃水浴中快速融化与在20℃水浴中中等速度融化效果相等,对菌的成活都没有太大影响。我们的试验肯定了在有保护剂存在下,用慢速冷冻可以在液氮中保存疫霉和霜疫霉。     

真鲷精子的超低温冻存及DNA损伤的检测

因名贵经济鱼类育种或种质资源保存的需要,精子超低温冻存及冻精质量评价的研究已越来越引起研究者的重视。在海产鱼类中,已见对大黄鱼(Pseudosciaena       

COMPARATIVE EVIDENCE FOR THE EVOLUTION OF SPERM SWIMMING SPEED BY SPERM COMPETITION AND FEMALE SPERM STORAGE DURATION IN PASSERINE BIRDS

Sperm swimming speed is an important determinant of male fertility and sperm competitiveness. Despite its fundamental biological importance, the underlying evolutionary processes affecting this male reproductive trait are poorly understood. Using a comparative approach in a phylogenetic framework, we tested the predictions that sperm swim faster with (1) increased risk of sperm competition, (2) shorter duration of female sperm storage, and (3) increased sperm length. We recorded sperm swimming speed in 42 North American and European free-living passerine bird species, representing 35 genera and 16 families. We found that sperm swimming speed was positively related to the frequency of extrapair paternity (a proxy for the risk of sperm competition) and negatively associated with clutch size (a proxy for the duration of female sperm storage). Sperm swimming speed was unrelated to sperm length, although sperm length also increased with the frequency of extrapair paternity. These results suggest that sperm swimming speed and sperm length are not closely associated traits and evolve independently in response to sperm competition in passerine birds. Our findings emphasize the significance of both sperm competition and female sperm storage duration as evolutionary forces driving sperm swimming speed.     

鲮鱼和二代混精鲮鱼低温耐受能力的差异

采用LKB循环冷却装置以程序降温方式比较鲮鱼及二代混精鲮鱼(草鱼混精鲮或多精混精鲮再用草鱼精液进行混精受精所得子代)的耐寒性能。无论是在7.0℃条件下的耐受时间,还是对低温耐受的极限,二代混精鲮鱼均优于鲮鱼,但这种改善是有限的。作者认为,混精受精法不能从根本上解决鲮鱼的不耐寒性能。应该寻求其他更有效的育种途径。     

昆明小鼠精子冷冻的研究(简报)

胚胎工程技术是动物品种、品系培育,种质资源保存及转基因动物制备、保种的重要手段。配子的冷冻保存技术目前广泛应用于胚胎工程。和胚胎冷冻相比小鼠精子冷冻技术方便、高效尤其适用于转基因及突变系小鼠的保种。成功的精子冷冻要求复苏后通过体外受精(IVF)获得胚胎,再移植入受体,生长发育并产出幼仔。胚胎体外培养获得足够的晚桑葚胚和早期囊胚是胚胎干细胞建立与制备嵌合体小鼠的成功关键,暂时不用的胚胎应仍能耐受冷冻保存,待复苏移植,建立新的繁殖群。但小鼠精子冷冻研究在我国开展的十分有限,缺乏相关资料数据。本实验对不同周龄SPF级昆明(KM)小鼠进行精子冷冻及冻融精子IVF,并将IVF获得的2-细胞胚胎分别进行胚胎移植、体外培养、胚胎冷冻及复苏后移植,应用冻融小鼠精子进行胚胎工程实践。     

水稻幼苗的低温伤害与膜脂过氧化

水稻幼苗在低温处理过程中,随着低温时间的延长和伤害的发展,叶片组织电阻急剧下降、膜保护酶超氧物歧化酶(SOD)活性降低、膜脂过氧化产物——丙二醛(MDA)含量明显增加。用四种自由基清除剂对水稻幼苗进行预处理,降低了低温引起的 MDA 含量的增生。相反,用 SOD 抑制剂预处理水稻幼苗,则提高了低温下 MDA 的含量。由此可以认为:水稻幼苗的低温伤害原因之一是与 SOD 活性受抑,加剧了自由基引发的膜脂过氧化作用,从而使膜受损伤有关。