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题名: 银鲫不同群体的分布格局、遗传多样性和系统关系研究
作者: 李风波
答辩日期: 2007-07-23
导师: 桂建芳
授予单位: 中国科学院水生生物研究所
授予地点: 水生生物研究所
学位: 博士
关键词: 银鲫 ; 转铁蛋白 ; 克隆多样性 ; 遗传多态性 ; 线粒体DNA ; 繁殖行为学
其他题名: Studies on distribution pattern, genetic diversity and phylogenetics among different populations of gibel carp, Carassius auratus gibelio
摘要: 银鲫(Carassius auratus gibelio)是一个能行雌核生殖的三倍体两性鲫鱼种群。以前关于银鲫的研究工作主要集中在二十年前从中国东北方正县双凤水库银鲫群体中鉴定出来的少数几个雌核生殖克隆上。尽管方正银鲫被保种和推广养殖多年以及在中国的几个其它地方也相继报道存在三倍体银鲫地方种群,但是我们对银鲫在天然水域的分布格局、养殖群体和自然群体中的遗传多样性状况及各群体之间的相互关系并不清楚,并且对其繁殖行为学研究也一直没有开展。本文通过野外广泛采样调查、流式细胞技术检测、几种分子标记技术应用和繁殖行为实验对以上问题进行了研究,并讨论了银鲫的地理分布特征、银鲫与二倍体鲫鱼共存的机制、银鲫克隆和遗传多样性产生的机制。 本文的工作主要分为四个方面: 第一,我们收集了来自新疆和长江中游的鲫属的14个野生群体,通过形态特征鉴定,确定这些群体均为鲫,没有发现黑鲫。随后,我们成功地应用流式细胞术对这些鲫鱼群体总共459个血细胞样品进行了倍性鉴定。通过调查发现,三倍体银鲫具有比以前想象的更广的地理分布,它出现在所有调查的水域;而二倍体鲫鱼仅出现于我们调查的长江中游水域。这些结果表明,银鲫存在“地理单性生殖”的现象,该现象在鲫鱼中是首次报道。基于银鲫群体存在少量的雄性个体(2.5 -16.7%),我们提出了“偶尔有性生殖”假说,认为银鲫偶尔进行有性生殖而产生少量的雄性个体,由此会付出一定的代价,因而可以部分弥补“性的双倍代价”,有助于银鲫与二倍体鲫鱼共存。我们也观察到,二倍体鲫鱼群体性比多数偏向雌性(1:2-1:8),意味着通过多产生雌性而减少“性的双倍代价”,也有利于银鲫与二倍体鲫鱼的共存,这一结果可能支持“偏向雌性的性别分配”假说。除了对于单性脊椎动物进化生态学研究的意义外,在不同水域发现的这些形态多样的三倍体银鲫将为银鲫优良品种的培育提供新的育种材料。 第二,我们用具有多态性的转铁蛋白标记对银鲫的不同地理群体进行了克隆鉴定。在取自黑龙江省的50尾方正银鲫个体中鉴定出10个不同的克隆,这一结果进一步被RAPD分子标记分析所证实;在来自河南省的51尾淇河鲫个体中鉴定出8个不同的克隆;从取自新疆的六个银鲫群体共133尾银鲫个体中鉴定出8个不同的克隆;从取自武汉长江江段的33尾银鲫中共鉴定出14个不同的克隆。由此,我们从调查的这9个银鲫群体的267个样品中总共鉴定出29个克隆,除了一个以前报道过的克隆A外,其它28个克隆是首次报道。因此,本研究结果支持这样一种观点,即单性脊椎动物群体中通常存在多个克隆。两个原种场(方正和淇河)中银鲫克隆多样性水平甚至高于一些银鲫野生群体,表明银鲫原种场中仍然保存了大量的银鲫克隆。我们推测有性生殖和重组是银鲫克隆多样性产生的主要原因。此外,如突变、父本遗传物质渗漏、多倍化、自然选择、杂交、迁移等因素也有助于克隆多样性的产生。我们发现克隆A是分布最广的克隆,这一现象可能支持“多面手基因型”模型,但也不能排除由于人类活动或者洪涝等自然灾害导致养殖池塘中的克隆A逃逸到了天然水体中所造成的影响。“冻结生态位差异模型”有助于解释不同银鲫群体多克隆共存的现象,但是还有待生态学研究来进一步证实。银鲫同时具有单性和有性的双重生殖方式可能是其克隆多样性形成和长期发展的生物学基础。在某些情况下,雌性银鲫可能摆脱对其有性亲缘种的精子依赖而选择其它鱼类的精子,既避免同有性宿主竞争又有利于占据新的生境,从而大大提高银鲫克隆的进化潜力。因此,这些新克隆的发现不仅在于其对进化遗传学研究的意义,而且有助于银鲫的进一步选择育种研究。 第三,为了获得银鲫遗传多样性和系谱关系的更多信息,我们从四个银鲫养殖场(方正、淇河、武汉和彭泽)中选取了64个有代表性的样品用于线粒体控制区(约920 bp)测序分析。这些样品代表了在养殖场中迄今发现的所有雌核生殖克隆。序列分析结果表明,遗传多样性水平在不同群体之间呈现显著差异,武汉和彭泽群体的几乎没有遗传变异,而方正和淇河群体的遗传多样性水平最高。系统发育分析表明,方正银鲫群体和淇河银鲫群体可能各自保存了一个不同的母系基因库。一个淇河银鲫样品含有方正群体的一个线粒体DNA单倍型,而广泛养殖的方正银鲫克隆A也拥有此单倍型。因此,这很可能是养殖的克隆A的个体逃逸到了淇河银鲫群体。四个方正银鲫样品聚到淇河银鲫样品形成的进化枝内,很可能反映存在从淇河群体到方正群体的基因流。分析结果也表明,武汉银鲫群体来源于方正银鲫群体,这与武汉群体从方正水库引种的事实是一致的。本研究结果显示,彭泽群体也来源于方正群体,为彭泽鲫起源于方正银鲫的假说提供了新的证据。 第四,我们观察并记录了银鲫雌雄鱼在室外的求偶行为,总结了银鲫的求偶通讯程序。观察发现,雌雄鱼婚配过程中,行为会自发协调一致,这将有助于提高其繁殖成功率。同时,也观察到银鲫在群水平下是一种混交方式。群体水平和个体水平雌性配偶选择实验表明,在模拟的自然条件下,银鲫雌性更倾向于选择同种的雄鱼,但是在少数情况下,它倾向于选择鲤鱼雄鱼。这种灵活多样的生殖行为将有助于银鲫产生遗传多样性和增强与有性亲缘种竞争的能力,对银鲫的长期进化具有重要意义。本研究的结论要推广到自然界中,还需要大量数据检验。
英文摘要: Gibel carp (Carassius auratus gibelio) is a triploid fish producing both gynogenetically and gonochoristically in China. Previous studies on it mainly focused on several limited clones identified twenty years ago in populations from Shuangfeng reservoir, Fangzheng County, northeast China. Although gibel carp has been preserved and popularized for years, and several triploid endemic populations of gibel carp have been also reported in China, little is known about patterns of distribution in natural water areas, the status of genetic diversity in cultivated and wild populations, and phylogenetic relationships among populations. Moreover, studies on reproductive behaviour have been never carried out in gibel carp from China. In this dissertation, these questions were addressed by field sampling across geographical regions, flow cytometry to detect ploidy level, combination of multiple molecular markers, as well as design of reproductive behaviour. Furthermore, several issues on evolutionary potential of gibel carp were discussed, including characteristic of geographical distribution, coexistence among different clones and with the sexual relatives, and mechanisms responsible for the origins of clonal and genetic diversity. This dissertation has been divided into four parts, as below: Firstly, fourteen wild populations of genus Carassius were collected from Xinjiang Province, northeast China and the middle reaches of the Yangtze River. After all the 459 samples were identified as C. aurassius based on morphological characters, their ploidy level was successfully determined by flow cytometry. Based on the result of ploidy analysis and field investigations, we found that triploid gibel carp was far more widely distributed than previously imagined, because it occured in all the sampling sites, most where gibel carp had never been reported; its closely related sexual, diploid C.a.auratus were found existing only in the middle reaches of the Yangtze River. In combination with previous, these results indicated that gibel carp shows a classic pattern of "geographical parthenogenesis", this phenomenon reported in C. auratus for the first time. Because of a few triploid fertile males (2.5 -16.7%) found in natural populations, here we propose the "occasional sex" hypothesis, holding that occasional sexual reproduction in gynogenetic gibel carp will produce a few males and thus partly compensates for the twofold disadvantage of sex, contributing to coexistence of gynogenetic triploid and sexual diploid C. auratus. Female-biased sex ratio (1 : 2 - 1 : 8) was observed in most of diploid populations, implying that the sexual form may mainly produce the female more than the male and thus the twofold cost of sex decreases. This process also promotes the coexistence of gynogenetic and sexual forms, supporting the "female-biased sex allocation" hypothesis. Besides the significance for evolutionary ecology of unsexual vertebrates, triploid gibel carp with diverse morphological characteristics discovered in different regions will be likely to provide new materials for selective breeding of excellent clones. Secondly, clonal diversity of gibel carp was examined in nine different geographical populations by using serum transferrin marker. Ten clones were clearly distinguished from 50 samples collected from Heilongjiang Province, further revealed by analysis of RAPD marker; eight, eight and fourteen clones were clearly identified in 51 samples from Henan Province, 133 from Xinjiang Province and 33 from Hubei Province, respectively. Therefore a total of 29 clones were detected in 267 samples in the nine populations, among which 28 were first found. This supports the notion that unisexual vertebrate populations are composed of numerous coexisting clonal lineages. Even, the level of clonal diversity in two national stock hatcheries was higher than that in some of wild populations, suggesting that they still preserve abundant clonal diversity. Sex and recombination may provide the principal source of clonal variation. Besides, other factors such as mutation, introgression of paternal DNA, polyploidy, natural selection, hybridization and migration also contribute to clonal diversity in gibel carp. The wide geographical distribution of clone A probably support the general-purpose genotype hypothesis, but we also can not exclude the possibility that clone A found in different sites is the one escaped from cultured ponds to natural rivers and lakes owing to the impacts of human activities and natural disasters such as flood. The frozen niche variation hypothesis is very helpful to explain coexistence of multiple clones, but further ecological studies are required to reveal it. The alternative reproduction modes of gynogenesis and sexual reproduction favor origin, maintenance and long-term evolution of clonal divisity. In some instances, females of gibel carp are likely to be independent of sperm of its closely related sexual males but select that of other fishes, thus avoiding competition with sexual hosts as well as facilitating colonization of new habitats. This change will greatly improve evolutionary potential of gibel carp clones. Furthermore, plenty of clones found will help to further selective breeding of gibel carp. Thirdly, to obtain more knowledge on genetic diversity and genealogical relationship within gibel carp, sixty-four samples representing all known clones from four hatcheries (FZ, QH, WH and PZ) were selected for complete mitochondrial DNA (mtDNA) control region (920 bp) sequencing. Genetic diversity varied remarkably among the hatcheries, with FZ and QH demonstrating high diversity and WH and PZ showing no variation. Sequence analyses indicate that FZ and QH might represent two distinct matrilineal sources. One of QH samples carried the haplotype shared by a most widely cultivated FZ clone, implying the probability the clone escaped from cultivated ponds to QH, whereas four samples in FZ clustered within the lineage formed mainly by QH samples, most likely reflecting historical gene flow from QH to FZ. Our data reveal that clones in WH originated from FZ, consistent with their introduction history, and support the hypothesis that gibel carp in PZ was actually domesticated from individuals of the FZ clone widely cultivated. Finally, after courtship behaviors in gibel carp were observed and documented outdoors, courtship communication were summarized. Synchronization of female producing eggs and males producing sperms will help to improve reproductive success. In a group mating experiment, females mate with males randomly. The following group and individual choice experiments indicated that in simulative natural conditions, females of gibel carp prefered conspecific males over males of carp (Cyprinus carpio), but occasionally the females demonstrated a preference for males of carp over conspecific. These flexible reproductive behaviors will facilitate origin of clonal variation and improving abilities of competition with closely related sexual species. If the results in this study are applied to natural conditions, much data are requred to test them.
语种: 中文
内容类型: 学位论文
URI标识: http://ir.ihb.ac.cn/handle/342005/12152
Appears in Collections:中科院水生所知识产出(2009年前)_学位论文

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银鲫不同群体的分布格局、遗传多样性和系统关系研究.李风波[d].中国科学院水生生物研究所,2007.20-25
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