银叶真藓适应极端环境的基因组特征及关键基因的作用机制

As the progenitors of terrestrial plants, bryophyte inhabitated on land about 4.7 hundred million years ago. They have evolved remarkable adaptability to cope with desiccation and extreme temperatures accompanied with dramatic environmental changes and widely distributed all over the world. Bryum argenteum Hedw. ( Bryaceae ) is a cosmopolitan unisexual species, which even survive extreme environments, such as the polar region, the equator, deserts and Tibetan Plateau. There are plenty of reports on the physiological and biochemical characters of B. argenteum in response to abiotic stresses. However, it will be difficult to address the underlining molecular mechanisms due to lack of genomic sequence information. We have collected nearly one thousand populations of B. argenteum from the different regions at different latitude, and set up the rapid clone regeneration system from a single gametophore to obtain homogeneous genetic background materials of each population. The genome size of B. argenteum is about 138 Mb measured by flow cytometry. We also have established technical platforms of protoplast regeneration, PEG mediated gene transformation and targeted gene knockout by homologous recombination. We are going to characterize the B. argenteum genome and screen for potential key genes involved in stress tolerance by de novo genomic sequencing, genomic resequencing, in-depth RNA sequencing and genome-wide association study (GWAS). Then we will verify the function of these candidate genes with molecular genetic, physiological, biochemical and cell biological tools. Our work will help to reveal the molecular mechanisms of the environmental adaptation of B. argenteum, and provide new gene resources for stress-resistance related plant genetic improvement.

约4.7亿年前陆地先锋植物苔藓登陆，在面临水分温度等迥异的环境胁迫下，藓类演化出极强的适应能力而分布广泛。银叶真藓是世界分布最广的物种之一，在地球两极、赤道、沙漠及青藏高原等极端恶劣环境中皆有分布。关于银叶真藓逆境应答的生理生化特征已有大量研究，但因无其全基因组序列，难于揭示其环境适应的分子机制。我们以纬度（年积温降水量）差异为准，采集近千不同居群材料，分别建立其单株“无菌快繁体系”，获得了遗传背景均一的纯系材料，细胞流式技术测定其全基因组为138Mb；建立了“原生质体再生、PEG介导基因转化、同源重组基因定点敲除”等技术。拟通过全基因组测序、重测序、深度RNA测序，以及全基因组与环境适应的关联分析（GWAS）等，分析银叶真藓适应极端环境的基因组特征，挖掘“关键基因”，并采用分子遗传、生理生化、细胞生物学等技术，验证基因功能，揭示银叶真藓环境适应的分子机制，为植物抗逆遗传改良提供基因资源。

聚焦于该项目“分析银叶真藓(Bryum argenteum)环境应答的基因组特征“，对其”关键基因“进行功能验证，揭示这种全球广布物种适应不同极端环境的遗传机制，为植物抗逆遗传改良提供基因资源，我们获得以下主要成果：.1.银叶真藓为雌雄异株植物。完成了雌株、雄株的全基因组序列分析，分别组装出高质量的11条染色体、注释出24,619和25,668个基因。.2.建立了真藓原生质体PEG转化系统，建立了同源重组基因敲除和CRISPR/Cas9基因编辑技术。.3.完成了漠河、芬兰、北京、宁波、舟山、云南建水等地的约600个居群材料（单株无菌纯系）的重测序。经过（正在进行中）GWAS分析，筛选出一个高温耐受相关转录因子ZFP1，采用基因同源重组和CRISPR/Cas9编辑技术获得相关突变体，研究结果显示，该基因与真藓感受高温环境密切相关，其分子调控机制正在进行中。.4.发现了控制性别的候选基因，建立了可控的“单倍体、二倍体完整植株”实验体系，正在进行的深入研究，可能在植物性别调控的分子机制领域取得“突破性成果”。.5.发表项目标注SCI论文2篇，待发表3篇；获批专利2个。.6.培养研究生6名（博士1名，硕士生5名）。