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杨属分子系统发育及三个山杨物种的生物地理学研究
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摘要
植物物种的系统发育关系和物种形成过程一直以来都是植物进化生物学研究中最基本的问题。系统发育关系的重建能够很好的解决各个物种间的关系,而物种形成研究是理解自然界物种多样性产生和变化的基础。准确估算各植物类群的出现时间对于推断其生物地理学具有非常重要的意义。在过去二十多年中,生物技术和理论方法的进步极大地推动了进化生物学的发展,促进了人们对一些重要植物类群系统发育关系、物种形成及生物地理学的认识,如拟南芥、玉米或水稻等。在木本植物中,利用核苷酸序列的变异式样对不同类群的系统发育关系、物种形成模式及生物地理学的研究已经非常成熟。但是对于模式植物杨属(Populus)来说,上述研究还鲜有报道。本文中,通过多基因位点序列的使用探讨了杨属分子系统发育关系及白杨派(section Populus)内中国山杨(P.davidiana)、欧洲山杨(P. tremula)和美洲山杨(P. tremuloides)的生物地理学。主要研究结果如下:
     1)使用生物信息学的方法,利用毛果杨(P. trichocarpa)和钻天柳(Salix arbutifolia)全基因组数据,筛选了24个在杨柳科内通用的单拷贝核基因位点作为遗传标记,并分别使用中国山杨和旱柳(S. matsudana)20个个体及杨属各派及柳属代表物种对其中15个位点的核苷酸多样性进行了计算。结果表明所有位点均含有丰富的系统发育信息并且均符合中性进化假设,满足了本文后续分析的需要。
     2)由于杨属种间杂交渐渗比较严重及种内形态变异比较大,杨属种间系统发育关系一直没有得到较好的解决。基于24个单拷贝核基因位点和12个叶绿体位点,对杨属各派及派内物种间系统发育关系做了深入研究。结果表明杨属是一个单系类群。在叶绿体系统发育树中,所有杨属物种形成两个分枝,一枝包括白杨派、大叶杨派及黑杨派和青杨派个别物种,另一枝包括胡杨派及黑杨派、青杨派的物种,这种拓扑结构表明杨属的母系祖先类群在形成初期即产生了分化。在核基因系统发育树中,胡杨派作为其他各派的基部类群,大叶杨派和白杨派次之,而青杨派和黑杨派占据系统发育树的末端位置。胡杨派与白杨派为单系类群而黑杨派和青杨派为复系类群。白杨派、黑杨派和青杨派很可能都是杂交起源的。对比分析核基因和叶绿体系统发育树可以发现,杨属中很多物种都是杂交起源的,如毛白杨、欧洲黑杨等。理解了这些物种的杂交起源方式有助于深刻剖析杨属中广泛存在的网状进化事件。在杨属的进化历史中,网状进化起到了非常关键的作用,同时也推动了杨属物种在不同分布区内的快速适应辐射。
     3)中国山杨、欧洲山杨和美洲山杨的外部形态非常相似并且亲缘关系非常近,这导致国内外学者对于三种山杨的分类学界定产生了矛盾,同时对于三种山杨的物种形成模式和生物地理学也缺少研究。本文中,通过大规模多居群多样本的采集,使用6个单拷贝核基因位点和3个叶绿体位点对上述问题进行了研究。结果表明,三种山杨都表现出非常高的核苷酸多样性,某些位点的多样性甚至与一年生植物相当。三种山杨之间产生了非常显著的遗传分化,Structure分析结果表明K=3为最佳聚类模型,并且三种山杨没有共享的叶绿体单倍型。上述结果表明将中国山杨划分为欧洲山杨的一个亚种或者将三种山杨合并为一个物种的分类学处理都是不合适的,三种山杨应该各自划分为独立的物种。叶绿体单倍型系统发育树和核基因位点物种树结果都表明美洲山杨是三个物种中最原始的类群。Beast宽松分子钟计算结果表明三个物种的分化处于第四纪初期。三个物种的祖先类群通过白令陆桥从北美大陆扩散至欧亚大陆并在较短时间内在欧亚大陆广泛分布。白令海峡的形成和青藏高原北部及邻近地区在第四纪的持续隆升将三个物种的分布区隔离开来。三个物种自共同祖先分化后种群经历了扩张-缩减的过程,并且彼此间并无显著的基因流,三个物种的物种形成模式为地理隔离形成导致的异域物种形成。
Inferring phylogeny and speciation process of plant species is one of the mostfundamental problems in plant evolution biology. Reconstruction of phylogeny will dissectrelationships among species and understanding speciation process is a prerequisite to explainorigin and variation of species diversity. It is of great helpful to infer the divergence and originof species accurately in detecting the biogeography history. In the past decades, advance onboth biological techniques and theoretical methodologies have remarkably reshaped ourunderstanding of phylogeny, speciation and biogeography history among model organisms,such as Arabidopsis, maize and rice. In tree species, researches on the phylogeny, speciationand biogeography history of different lineages using the variance pattern of nucleotidesequence have accumulated, however, rareness has been reported in model tree species,Populus. In this study, using multiple nucleotide loci, the molecular phylogeny of Populus andthe speciation model and biogeography of three aspens in section Populus, P. davidiana, P.tremula and P. tremuloides were explored. The main results are as follows:
     1)24single-copy nuclear markers were developed through bioinformatics methods basedon whole genomic sequences of P. trichocarpa and Salix arbutifolia. Twenty individuals of P.davidiana and S. matsudana were used to evaluate the basic application of15loci with respectto marker length and diversity indices, respectively. Results indicated that all the15locicontained abundant high phylogenetic information and fitted the null hypothesis of neutralevolution well. The utility of single-copy nuclear markers will facilitate subsequent studies onphylogeny, population genetics and biogeography, as well as the evolutionary dynamics ofSalicaceae.
     2) The complex reticulate evolution (hybridization, introgression) and highly variableintraspecific morphological traits have posed difficulties in resolving the phylogeny of Populus.Based on a large data set of nuclear and chloroplast DNA sequences, we reconstructed robustphylogeny of Populus using maximum parsimony, maximum likelihood and Bayesian inference methods. The resulting phylogenetic trees showed better resolution at both inter-andintra-sectional level than previous studies. The results revealed monophyly of Populus.CpDNA based phylogenetic tree resulted in two main clades, one consisted section Populus,Leucoides and species of section Aigeiros and Tacamahaca, the other comprised of sectionTuranga and the remaining species of section Aigeiros and Tacamahaca. This topologysuggested an early divergence of the maternal progenitors of Populus. In the nuclear DNAbased phylogenetic tree, section Turanga occupied the basal position following sectionPopulus and Leucoides, section Aigeiros and Tacamahaca took the terminal position. SectionPopulus and Turanga were monophyly and section Aigeiros and Tacamahaca were polyphyly.The three advanced sections, Populus, Aigeiros and Tacamahaca may be hybridized origin.Comparative analysis of the cpDNA and nuclear DNA based phylogeny elucidated that somespecies were hybrid origin, such as P. tomentosa, P. nigra and P. deltoides. Realizing the hybridorigin of species in Populus will conspicuously promote our understanding the reticulateevolution in Populus, which may have played a significant role in the evolution history ofPopulus by facilitating rapid adaptive radiations into different environments.
     3) Morphological traits and relationships among P. davidiana, P. tremula and P.tremuloides were close, which caused huge controversy on the taxonomical delimitation of thethree aspens. Furthermore, speciation model and biogeography of these three aspens havenever been reported before. In this study, these questions were addressed using multilocusnucleotide sequences and multiple samples. All the three aspens showed high nucleotidediversity, which was comparable with that of annual plant species in some loci. Significantgenetic differentiation was detected among the three aspens and Structure results showed thebest clustering model when K=3. Furthermore, no shared cpDNA haplotype was probed. Basedon these findings, we proposed that previous taxonomic classification which classified P.davidiana as a subspecies of P. tremula or classified these three aspens as one species wasinappropriate. The three aspens should be delimitated as three single species. Phylogenetic treebased on cpDNA haplotypes and BEST species tree based on6nuclear loci showed that P.tremuloides was the more ancestral lineage in the three species. Divergence of the three species inferred from Beast relaxed molecular clock occurred in early Quaternary. The ancestrallineage dispersed from North America to Eurasia through Beringia land bridge (BLB) followedby fast expansion throughout Eurasia. The formation of Bering strait and the uplift of thenorthern Qinghai-Tibetan Plateau (QTP) and adjacent area in Quaternary isolated thedistribution of the three species, which have undergone a process of populationexpansion-contraction dynamics since divergence from the common ancestor. No obvious geneflow was detected which suggested that the present patterns of nucleotide variation was notshaped by migration among these species and that introgression was not pervasive in the groupof species. The speciation model of the three species was allopatric speciation caused byvicariance.
引文
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