桑树绿枝扦插皮部生根的生理与miRNAs调控机制研究

Cuttage is an effective method for asexual propagation in woody plants. Adventitious root (AR) formation is one of the key factors for a successful cuttage. However, the physiological and molecular mechanisms of rooting in mulberry cuttings remain unclear. The new propagation technique developed by our research group can provide ready access to large amount of high quality cutting materials, making it possible to comprehensively investigate the mechanisms of AR formation in mulberry cuttings. Basal bark of the softwood cuttings from mulberry cultivars with contrasting rooting ability are selected to analyze the dynamic changes of physiological parameters including plant phytohormones, nutrients, ROS (reactive oxygen species) and rooting-related enzymes in different rooting stages. Furthermore, by using high-throughput sequencing, we intend to screen the key miRNAs which may link closely to the AR formation in mulberry cuttings, and to identify their biological functions through transgenic technology. Combining the physiological and molecular analysis mentioned above, we aim to uncover the physiological and miRNAs-directed regulational mechanisms of cuttage rooting in mulberry. This study has significant theoretical and practical implications. It will not only enrich and develop our understandings to the regulation pattern of rooting in mulberry cutting, but also cast a light on the technique innovation of cutting nursery in other hard-to-root plant species.

扦插育苗是木本植物无性繁殖的有效手段，不定根的形成是植物扦插繁殖成功的关键，而有关桑树扦插生根的生理基础和分子机制并不清楚。本研究组建立的桑树扦插快繁技术为全面地分析桑树扦插生根机理提供了高质量的试验材料。本项目拟以绿枝扦插生根能力显著不同的桑树品种为研究材料，从生理生化水平，分析它们在不定根发育的不同阶段植物激素、营养物质、活性氧ROS以及生根相关酶类等的动态变化规律；应用高通量测序技术，筛选出与桑树扦插生根密切相关的miRNA基因；应用转基因技术，鉴定它们在生根过程中的生物学作用。综合以上结果，揭示桑树扦插不定根发育的生理与miRNA调控机制。本项目不仅是对桑树扦插根发育调控模式的丰富和发展，也为其他植物扦插育苗技术的创新提供理论指导，具有重要理论和实践意义。

不定根的形成是植物扦插繁育成功的关键环节。通过人工诱导，桑树插穗扦插不定根可从皮层薄壁细胞直接增殖分化形成根原基。然而，这种高效的皮部生根形式的调控机制还不清楚。本项目以湘7920和育711两种生根能力不同的桑品种为主要研究对象，首先比较了皮部生根起始期、膨大期和伸长期等三个不同发育阶段基部皮层的基因转录表达差异，发现膨大期时插穗的转录水平变化最显著；基于该研究结果，利用生理生化技术，剖析了皮部生根初期，即根原基诱导过程中的植物激素、营养物质、活性氧以及生根相关酶类等的动态变化规律，发现皮层中瞬时高浓度生长素可能是促进根原基分化的化学信号之一。转录组分析发现生物节律、植物-病原互作、蔗糖代谢、胁迫响应等通路也在不定根原基诱导中的重要作用。另一方面，应用RNA-seq技术和关联分析，鉴定了皮部生根和常规生根不定根不同发育时间皮层中的差异表达基因及miRNA，筛选到若干受吲哚丁酸诱导的miRNA-mRNA对，揭示了激素诱导的桑树扦插皮部生根的转录水平调控机制。研究结果为阐明植物扦插不定根发生提供了新的试验依据，也为其他难扦插生根植物的繁育技术创新提供了理论指导。