基于EST-SSR变异的青藏扁蓿豆(Medicago archiducis-nicolai)环境适应机制解析

Medicago archiducis-nicolai Sirjaev is an allogamous diploid perennial species and one of the cohesive core in the section Platycarpea of the genus Medicago. It appears to be one of remnants of an ancient Arcto-Tertiary complex of flora forebearing many new genera including Medicago and Trigonelle. It has been found that M. arciducis-nicolai is widely distributed in the Qinghai-Tibet Plateau (QTP) and its adjacent areas as the only species in the genus Medicago that can survives the extremely harsh environments mainly caused by altitudinal difference in the QTP, M. archiducis-nicolai is an ideal species for elucidating of the mechanisms of plant adaptation to the extreme environment in the QTP. In an attempt to dissecting the molecular genetic mechanism of M. archiducis-nicolai adaptation to the extremely harsh environments in the QTP and casting light on improvement for abiotic stress tolerance in the cultivated species of the genus Medicago, especially for winter hardiness, this study plans to develop abiotic stress- and development-related EST-SSR markers based on our now available RNA-Seq data of cold treated M. archiducis-nicolai, and to search for molecular signatures of divergent selection in M. archiducis-nicolai natural populations by identification of EST-SSR outlier loci associated with the environmental factors. EST-SSR alleles associated with climatic variables will be investigated to reveal genetic polymorphisms contributing to local adaptation on the QTP. Moreover, the roles of candidate genes revealed by potential divergent selected EST-SSR loci will be further verified taking advantages of known knowledge and validated by experiments.

青藏扁蓿豆(Medicago archiducis-nicolai)为苜蓿属（Medicago）阔荚苜蓿组(Section Platycarpae)的核心种之一，属古北极第三纪植物区系残遗种(remnant)，广布于青藏高原及其毗邻地区，对海拔差异导致的局部异质性极端生境具有极强的适应性，是研究青藏高原极端环境植物适应机制的理想物种。本项目拟利用已经获得的低温胁迫转录组测序信息，开发青藏扁蓿豆其逆境胁迫和发育相关基因的EST-SSR标记；在中性遗传标记群体遗传多样性和遗传结构分析基础上，鉴定环境选择相关EST-SSR离群位点(outlier loci)，分析气候因子与等位基因频率的关联性，筛选与极端环境适应相关的基因组区段或基因；进行部分候选基因的生物学功能解释和实验验证。希望通过上述研究工作，部分解释青藏扁蓿豆适应青藏高原极端环境的分子机制，也为栽培苜蓿抗寒性定向改良提供参考信息。

青藏扁蓿豆(Medicago archiducis-nicolai)为苜蓿属（Medicago）阔荚苜蓿组(Section Platycarpae)的核心种之一，广布于青藏高原 及其毗邻地区，对海拔差异导致的局部异质性极端生境具有极强的适应性，是研究青藏高原极端环境植物适应机制的理想物种。本项目利用转录组测序信息，设计开发了青藏扁蓿豆EST-SSR标记。验证了引物在青藏扁蓿豆基因组中扩增的有效性。利用开发的EST-SSR标记对青藏扁蓿豆和扁蓿豆的中性群体遗传结构和遗传多样性进行了分析。EST-SSR标记可以清晰区分青藏扁蓿豆和扁蓿豆两个物种；青藏扁蓿豆群体间存在明显的遗传分化，形成了高原台面和东部台面与边缘分布的两个组群；青藏扁蓿豆群体间遗传分化与地理距离、纬度、海拔高度差之间存在极显著的相关性，表明地理距离和青藏高原复杂的地貌环境导致了青藏扁蓿豆群体间的遗传分化。青藏扁蓿豆群体遗传多样性普遍低于扁蓿豆，符合狭域分布物种群体的遗传多样性特点；位于高原腹地的青藏扁蓿豆群体遗传多样性高于分布于边缘的群体，提示在高原腹地存在潜在的避难所。对青藏扁蓿豆群体中EST-SSR离群位点检测，获得了7个和8个分别受到正向选择和平衡选择的位点。对上述候选离群位点结合气候变量空间分析，以及单倍型基因频率与气候变量之间的回归分析，获得了涉及3个位点的3个单倍型变异。上述与气候因子显著相关的3个单倍型变异所涉及的基因与开花时间调节、线粒体磷酸根转运和RNA加工和稳定性调控有关。通过本项目所获得的研究结果为青藏扁蓿豆遗传资源的保护和利用、解释适应极端环境的机制，以及苜蓿属栽培种的抗逆性改良提供了参考信息。