蓖麻矮化相关RcDof基因功能分析及调控机制研究

Castor (Ricinus communis) is a typical oilseed crop which is widely cultivated in Inner Mongolia. The dwarf castor has many advantages, such as high yield, high density planting and mechanization, etc. Although breeding in dwarf castor has been studied for many years, the mechanics of transcription in dwarf gene is unclear. The Dof (DNA-binding with one finger)domain proteins are plant-specific transcription factors with highly conserved DNA-binding domain, and play critical roles as transcriptional regulators in plant growth and development, including regulating plant dwarfing. In our research, a Dof gene (RcDof gene) of zinc finger protein was found that involved in the process of castor dwarfing. The purpose of this study is to clone RcDof gene, transform it into high stalk castor and analyse its function. According to determining the target genes, regulating pathways and protein-protein interaction, we can study its regulatory mechnism of dwarf. Our research will provide new germ plasma resources, and contribute to improving the yield of castor.

蓖麻是内蒙古地区具有区域特色的油料作物，矮秆蓖麻品种具有单产较高、适合密植和机械化生产等优点。蓖麻矮化育种研究开展多年，但矮秆基因的转录调控机制及作用机理尚不清楚。Dof(DNA-binding with one finger)蛋白是植物特有的一类转录因子，在植物生长发育过程中起着重要的作用，可以调控植株矮化。本课题组在以往的研究中发现了一个锌指蛋白RcDof基因，该基因在矮秆蓖麻材料中表达下调。因此，本研究拟克隆蓖麻锌指蛋白RcDof基因，转化高秆蓖麻材料，分析该基因的功能；明确RcDof基因作用的靶基因、调节途径及与RcDof基因编码蛋白相互作用的蛋白质，探讨其矮化调节机制,为蓖麻矮化育种提供新材料，为提高蓖麻单产奠定基础。

蓖麻是内蒙古地区具有区域特色的油料作物，矮秆蓖麻品种具有单产较高、适合密植和机械化生产等优点。蓖麻矮化育种研究开展多年，但矮秆基因的转录调控机制及作用机理尚不清楚。Dof(DNA-binding with one finger)蛋白是植物特有的一类转录因子，在植物生长发育过程中起着重要的作用，可以调控植株矮化。本研究根据NCBI发表的基因序列，设计引物并克隆了蓖麻锌指蛋白RcDof基因，构建了RcDof基因过表达及基因沉默表达载体；采用农杆菌介导法转化高秆蓖麻材料通蓖5号，经抗性筛选及转基因植株分子生物学鉴定，获得了转基因矮化植株；转基因植株与野生型植株表型分析显示，RcDof基因过表达植株变矮，而RcDof基因表达量下降植株较高，证实了RcDof基因参与了蓖麻株高的形成。转基因植株与野生型进行转录组学分析，筛选差异表达基因1959个，其中上调基因1201个，下调基因758个。KEGG通路富集分析显示，植物激素信号通路中有22个基因差异表达，通过qRT-PCR定量分析验证生长素相关通路基因，明确RcDof基因主要影响了生长素代谢途径进而影响株高。亚细胞定位结果显示，RcDof基因定位于细胞核内。构建了RcDof基因大肠杆菌表达系统，结果显示全长蛋白以包涵体形式表达。制备了大量RcDof（44-101）截短体蛋白，并偿试筛选与RcDof相互作用蛋白。所得结果为蓖麻矮化育种提供新的视角及新的种质资源。