Species delimitation in the apomictic polyploid Ranunculus auricomus complex using an integrative TaxonOMICs approach

在无融合生殖植物中，物种的界定仍然是一个挑战。通过种子进行无性繁殖。无融合生殖多倍体复合体通常包括一些有性祖先物种和许多无融合生殖杂种衍生物。兼性性行为可以导致形成许多局部分布的形态表型。特别是在北方欧洲，许多这样的形态型传统上被描述为无性种，而不考虑任何进化或遗传背景。为了克服这种对毫无根据的实体的纯粹描述性分类，该项目的目标是根据在空间和时间上具有长期持续性的进化谱系来界定物种。我们将完成对北方欧洲的欧亚毛茛复合体（超过600种已描述的形态种）的分析，通过使用综合方法结合现有方法：（1）系统基因组学，结合两种下一代测序方法，即：核基因的靶向富集与质体基因组略读相结合，用于基于聚结的物种划界和限制性位点相关DNA测序（RAD-seq），作为评估物种内和物种间多样性的精细尺度基因组标记。目标富集的一个子集的c。600个基因的核基因将解决分类群的系统发育框架，并将用于研究空间和时间的多样性与地理分析和分子测年。RAD-seq将用于分析起源后进化，基因流和无融合生殖谱系内和之间的多样性。(2)将使用流式细胞术通过流式细胞术种子筛选（FCSS）确定倍性水平和兼性程度。(3)植物的数字成像、几何形态测量学和多元统计分析将用于客观地评估分类群之间的形态分化和分类群内的多样性。分类将基于结果的组合，以正式地将仅确定的进化单位识别为物种。与其他项目的合作包括为无性生物和植物多倍体复合体的分类以及方法方面制定一个全面的物种概念。

Species delimitation is still a challenge in plants with apomixis, ie. the asexual reproduction via seeds. Apomictic polyploid complexes usually comprise a few sexual progenitor species, and numerous apomictic hybrid derivatives. Facultative sexuality can result in the formation of numerous, locally distributed morphological phenotypes. Particularly in Northern Europe, numerous such morphotypes have been traditionally described as agamospecies without regarding any evolutionary or genetic background. To overcome this solely descriptive classification of ill-founded entities, the project is aiming at a species delimitation based on evolutionary lineages that have a long-term persistence in space and time. We will complete the analysis of the Eurasian Ranunculus auricomus complex in Northern Europe (>600 described morphospecies) by using an integrative approach combining established methods: (1) Phylogenomics, with a combination of two next-generation sequencing methods, ie. target enrichment of nuclear genes combined to plastid genome skimming for coalescence-based species delimitation and restriction site-associated DNA sequencing (RAD-seq) as a fine-scale genomic marker to assess diversity within and among species. Target enrichment of a subset of c. 600 genes nuclear genes will resolve the phylogenetic framework of taxa, and will be used to study spatial and temporal diversification with biogeographical analysis and molecular dating. RAD-seq will be used to analyze post-origin evolution, gene flow and diversity within and among apomictic lineages. (2) Flow cytometry will be used to determine ploidy levels and degree of facultative sexuality via flow cytometric seed screening (FCSS). (3) Digital imaging of plants, geometric morphometrics and multivariate statistical analysis will be used to objectively assess morphological differentiation among taxa and diversity within taxa. Classification will be based on a combination of results to recognize formally only established evolutionary units as species. Collaborations with other projects include the development of a comprehensive species concept for classification of asexual organisms and for plant polyploid complexes, and on methodical aspects.