海带、裙带菜和铜藻的基础繁育生物学研究及产业化应用

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题名	海带、裙带菜和铜藻的基础繁育生物学研究及产业化应用
作者	李静
学位类别	博士
答辩日期	2015-05-15
授予单位	中国科学院大学
授予地点	北京
导师	逄少军
学位专业	海洋生物学
中文摘要	海带、裙带菜及铜藻是我国重要的经济褐藻，开展这三种海藻的基础繁育生物学研究，了解并掌握它们的繁育特征和生殖规律，可以指导我们建立开发和利用它们的技术方法，这对于解决育种产业中的现实问题具有重要的意义。本论文在实验室及养殖海域条件下，开展了海带、裙带菜精子寿命、裙带菜雌雄同体配子体生活史及铜藻受精卵低温保存等研究，具体研究内容和结果如下： 1）观察统计了海带、裙带菜卵子的排放规律，并首次采用统计受精率的方法对海带、裙带菜精子寿命进行了测量。得到裙带菜精子在18℃黑暗条件下仅能存活1小时，而海带精子在12℃黑暗条件下能存活12小时以上的结论。随后结合生产化养殖模式，以SSR分子标记为手段证实了海带夏苗生产化工艺中的确存在精子污染、品种混杂的现象。依据研究结果，在育苗企业实践了单独的海带亲本维护系统，从而在技术和工艺层面上解决了大生产过程中品种混杂的问题。 2）通过对裙带菜雄配子体的单独培养，发现了雌雄同体现象的存在，并对这一雌雄同体配子体展开了深入的研究。在三个裙带菜养殖品系中，总共统计得出43%的配子体出现了雌雄同体现象，说明了这一现象是普遍存在的。针对这一配子体，我们在实验室中得到了它的自交孢子体，并在海区中培育至性成熟采集了游孢子，游孢子萌发之后除了15-20%的配子体形成雌雄同体结构外，其他全部发育成雄配子体。而且，采用与雌性相关联的微卫星标记对雌雄同体配子体、自交孢子体、以及自交孢子体释放的游孢子发育得到的子代配子体进行检测，发现它们全部缺失与雌性相关联的条带。同时，利用现有的20对裙带菜微卫星引物对三者进行检测，得到的等位基因片段长度完全一致。根据孤雌生殖全部释放雌性游孢子的特点，再结合自交孢子体全部释放雄性游孢子，提出了雌雄游孢子杂交育苗这一新的技术手段，并成功将这一技术应用到了生产实践中。 3）采用实验生物学的方法，探讨了铜藻生殖托上自然脱落的受精卵经过4℃的短期保存，受精卵的成活及后期的发育情况。结果显示，4℃低温条件下培养6小时和20小时的受精卵与相同时间15℃适温条件下培养的受精卵，死亡率无明显差别。受精卵经过最长20小时的低温保存后恢复到适温条件下培养，与一直培养在适温条件下的受精卵相比，后期幼苗的形成和附着力无显著差异。这一研究结果为跨地域运输铜藻受精卵，实现铜藻的异地育苗提供了依据。
英文摘要	Sacharina japonica, Undaria pinnatifida and Sargassum horneri are three important economic seaweeds in China, study on the basic reproduction biology of the three species and document their reproduction characteristics can help us improve the breeding technology and solve the aquaculture problems. The thesis includes the evaluation of the sperm life-span of S. japoncia and U. pinnatifida, the life-cycle of monoecious gametophtyes of U. pinnatifida and the cryopreservation of zygotes of S. horneri. 1）The potential life-span of the sperm of two seaweeds, S. japonica and U. pinnatifida, was first assessed by their ability to fertilize eggs. Results show that some sperm of S. japonica stored in dark at 12°C were able to fertilize eggs up to 12 h after discharge, while the sperm of U.pinnatifida can only survive 1 h in the dark at 18℃. Sperm-contamination was further confirmed in large-scale breeding of S.japonica based on microsatellites. According to the results, independent cultured system for parental-sporophytes was established to solve the mixture problem in the breeding company. 2）The unusual zoospore-derived monoecious gametophytes of U.pinnatifida were first discovered. To understand this phenomenon, a comprehensive screening in previously identified male gametophyte cultures isolates was carried out. Forty-three percent of the isolates developed both antheridia and oogonia on the same filamentous fragment, suggesting that the phenomenon was common in the male gametophytes of the cultivars tested. Selfing of the monoecious gametophyte gave rise to morphologically normal sporophytic offspring. The selfed sporophytic offspring were grown to maturity in open sea cultivation. Active zoospores were released from mature sporophylls. The majority of these developed into male gametophytes, while 15–20% developed into the observed monoecious structures on the same filament. Using PCR amplification it was found that all the monoecious gametophyte isolates, the selfed sporophytic offspring and the F1 gametophytes derived from zoospores released by the selfed sporophytes lacked the female-linked microsatellite sequence (a part of the locus Up-AC-2A8, GenBank accession No. AY738602.1), indicating their male nature. Meanwhile, they also have the same genotypes at twenty SSR loci. With the new discovery of the male sporophyte obtained by selfing the monoecious gametophyte, in combination with the previous finding of the potential of parthenosporophyte to release female spores, crossing of the spores from these two lines of sporophytes was successfully applied to the seedling production. The finding of this investigation will pave a way of using pure sporophytic lines to cross for selecting offspring with expected agronomical features demanded by the farming industry in the future. 3）The development of zygotes naturally dropping from the conceptacles of S. horneri were investigated after stored at 4°C for fixed time periods. The results showed that the zygotes mortalities conserved at 4 °C for 6 and 20 hours were the same as that of the zygotes preserved at 15°C. The zygotes cultured at 4°C for less than 20 hours showed no problem to attach and grow on the substratum. Discoveries of this investigation provided data support of transporting the zygotes over distance under low temperature for seedling production when necessary.
公开日期	2015-06-11
内容类型	学位论文
源URL	[http://ir.qdio.ac.cn/handle/337002/23255]
专题	海洋研究所_实验海洋生物学重点实验室
推荐引用方式 GB/T 7714	李静. 海带、裙带菜和铜藻的基础繁育生物学研究及产业化应用[D]. 北京. 中国科学院大学. 2015.