水杉花芽调控分子机制的研究

Trees with genetic dwarf traits can easily bear flowers and seeds, but its seeds are of low-quality. For fast-growing superior strains, due to their fast-growing physiological mechanisms inhibiting the flower bud differentiation,it is difficult to bear fruits. And that is why forest seed orchard at home and abroad for decades failed to meet the seed yield. What’s more, that’s the main reason to limit the process of afforestation, especially more difficult for gymnosperms. At present，little is known about the molecular mechanism in woody plant species, especially in gymnosperms. We have studied for many years about a technology that induces cone formation. Now on this basis,we can handle the physiological process of the flower bud differentiation, analysis the transcriptome responses in flowering phase using 454 pyrosequencing,analysis the regulation of a series of related genes expression of Metasequoia in different ages and to the point,unrevel the molacular mechanism of reproduction initiation in Metasequoia. The current study not only provide the theory of physiological and molecular regulation in the exploration of tree breeding but also establish a solid foundation for the population protection and development of the relict Metasequoia. In addition，it lay the theoretical foundation on the technological innovation of all kinds of fast-growing tree seed orchard.

遗传矮小性状的林木极易开花结实，其种子通常劣质，而速生优株因速生生理机制抑制其花芽分化难以结实，这就是国内外林木种子园数十年来也未能达到良种丰产的目的的原因，也是限制造林良种化进程的主要原因。特别是裸子植物开花结实的难度更大。目前，林木植物，特别是裸子植物花芽分化的分子调控机制的认识还非常有限。我们已研究多年人工促进水杉雌花形成花球并结实的技术，在此基础上通过人工调控不同阶段水杉花芽分化的生理进程，利用454高通量测序技术对花芽分化阶段的转录组进行测序，分离鉴定与水杉开花调控和植物激素相关的基因并分析其功能，揭示水杉开花调控分子机制。该研究是对林木育种生理、分子调控理论的探索，为濒危孑遗树种水杉的种群保护和推广栽植奠定科学依据，并为各种林木种子园速生丰产的种子的技术创新奠定理论基础。

项目以水杉作为研究材料，使用通过454高通量测序技术对水杉的营养芽和雌球花芽进行转录组测序，构建了水杉的EST数据库，共得到97,565条高质量水杉转录本序列；与公用数据库进行了比较分析，注释和分类，共检测到59个与开花相关的基因家族和190个参与激素调控的转录本。同时通过Illumina测序建立了水杉雌雄花芽分化时期的表达谱，共检测到69,520 (71.3%)个差异表达的基因，发现了参与雌雄花芽分化过程及糖代谢途径、激素调控中表达丰度显著变化的基因。项目筛选了水杉荧光定量PCR中稳定表达的内参基因，克隆到了水杉激素处理后花芽分化及花器官形成过程中的差异表达基因，包括2个LFY（LEAFY）家族成员、3个UFO基因家族成员及14个MADS-box家族成员。将MgLFY和MgNLY基因在转基因拟南芥的超表达，发现35S::MgLFY和35S::MgNLY转基因拟南芥株系开花的时间早于野生型Col拟南芥，MgLFY、MgNLY可以诱导拟南芥提前开花，从而缩短成花周期。 研究结果为研究水杉花芽分化及花器官形成及裸子植物花发育分子机制提供了丰富的基础数据，为濒危孑遗树种水杉的种群保护和推广栽植提供科学依据。