基于RAD tags方法的生物地理学研究：解析东亚岛屿特有植物舟山新木姜子地理分布格局形成与谱系分化机制

Quaternary climatic oscillations have caused changes in sea level that altered the size, number and degree of isolation of islands, particularly in continental-shelf islands. Elucidating the demographic effects of these oscillations and environmental heterogeneity increases our understanding of the roles of climate change and niche divergence in shaping biodiversity in East Asia. In this study, we will study on the lineage divergence, geographic pattern and genetic diversity of Neolitsea sericea, endemic to East Asian continental-shelf islands. The samples of Neolitsea sericea have been sampled from different islands across the distribution range of the species in China, Japan and Korea. The sequence variation of both chloroplast DNA (cpDNA) and nuclear ITS as well as nuclear SSR dataset have been used to investigate the geographic distribution patterns, phylogeographic structure, and phylogenetic relationships of haplotypes. Based on the patterns of lineage diversification, we further use a genome-wide sampling of SNPs identified within restriction site-associated DNA (RAD) tags to investigate the historical forces responsible for geographic pattern and the mechanisms of lineage diversification. The population genetics methods and Bayesian and ABC models will be used to infer demorgraphic history of species and lineages. When combining with ecological niche modeling and multivariate niche method, we can address what historical processes, e.g. fragmentation, expansion, long-distance colonization, have the strongest influence on the lineage divergence and geographic pattern of N. sericea. By using genome scan methods, we can identify RAD neutral and outlier loci. Based on landscape genetics methods, we will model environmental data using nonparametric multiple regression to explain lineage divergence and genetic differentiation as a function of spatio-ecological effects. In addition, to identify patterns of adaptive genetic variation on landscapes, and quantify effects of environmental factors on patterns of adaptive genetic variation, we will test the correlations between clines in allele frequencies of individual outlier loci and environmental variation. Based on multi-loci genetic data, evolutionarily important units can be identified. This information will be useful for developing scientific conservation and appropriate management strategies for this endangered species.

揭示气候动荡与环境异质性对大陆架岛屿生物类群种群动态的影响，有利于理解东亚物种多样性形成机制。本项目拟对东亚岛屿特有植物舟山新木姜子进行广泛群体取样，在叶绿体、ITS，nSSR分析的谱系地理结构基础上，应用限制性内切位点相关的DNA标记（RAD tags）技术鉴定群体单核苷酸多态性（SNPs）。基于整合数据，利用群体遗传学、ABC模型方法分析物种及谱系的进化历史。利用Bayesian方法分析谱系分化过程，结合生态位模型与多变量生态位方法，揭示谱系分化的历史驱动因子。通过基因组扫描方法鉴定RAD的中性与outlier位点，利用景观遗传学方法分析地理与景观效应、生态因子对群体/谱系分化以及遗传多样性空间格局的影响，解析适应性遗传变异的空间分布及其影响因子。中性与适应性位点结合鉴定"进化重要单位"，提出濒危植物舟山新木姜子遗传资源保护与管理策略。

本课题对东亚岛屿分布的特有植物舟山新木姜子进行了广泛的野外资源调查与取样，基于叶绿体与核基因组序列、微卫星以及简化基因组数据，利用亲缘地理学以及群体遗传学方法研究了舟山新木姜子的群体遗传多样性、谱系地理结构、基因流以及群体进化历史，并利用物种分布模型、景观基因组等方法分析了其谱系分化的驱动力。获得如下研究结果：.（1）.野生资源状况与遗传多样性。野外调查表明，舟山新木姜子主要分布中国东部舟山群岛与台湾、韩国南部、日本本岛南部以及琉球群岛，约60多个群体，采集61个群体DNA样本进行遗传多样性分析。基于群体水平的ITS 序列，获得了3 个ITS 单倍型。基于cpDNA（psbA-trnH、trnL-trnF、psbC-trnS、rps16）序列，获得20 个cpDNA 单倍型，微卫星（EST-SSR）与叶绿体序列水平的群体遗传多样性随纬度的升高而降低。.（2）.谱系地理结构。基于叶绿体、ITS、微卫星以及简化基因组信息(RAD-seq)的群体结构分析显示，舟山新木姜子以Tokara Gap为界分化成南北两个谱系，北部以濑户内海为界的两个亚谱系。基于RAD-seq数据，以最大似然法构建的系统发育树揭示了舟山新木姜子在地质历史时期经历了由南向北的迁移事件。.（3）.谱系分化与群体动态历史：基于微卫星及RAD-seq数据利用近似贝叶斯（ABC）及随机森（Random Forest）等多种拟合分析表明，南北谱系分化时间为0.33 Ma，北部亚谱系分化时间为0.036 Ma。南部谱系在0.026 Ma经历了瓶颈事件，南北谱系间不存在近代基因流。根据谱系分化的估算时间以及古地质、气候资料证据表明，更新世中晚期以来Tokara地区陆桥下沉是南北谱系分化的根本原因，而末次盛冰期“避难所”则导致了北部次级谱系的分化，而冰后期海平面上升则导致目前中国、日本本岛南部、韩国的舟山新木姜子群体间断分布式样。.（4）.谱系分化的驱动力：景观基因组学研究表明，除地理与景观的阻隔效应外，‘Tokara gap’南北两侧的环境因子尤其是温度因子差异所产生的自然选择在舟山新木姜子的谱系分化过程中也起到了非常重要的作用。. 最后，根据遗传资源与生态位模拟的研究结果，我们提出了舟山新木姜子科学保护的策略。