PtrFBL7基因在杨树次生生长中的功能研究

Auxin plays an important regulatory role in plant growth and development via its signal transduction. Auxin receptor is one of the most critical components in auxin signaling pathway. In the previously study, analysis of transport inhibitor response 1 (TIR1) gene family in Populus (designed as PtrFBLs) was performed, including sequence analysis, structural properties, phylogeny and expression patterns. PtrFBL7 was mainly expressed in vascular and cambial tissues, indicating that its function was involved in secondary growth. To reveal the role and regulation mechanism of PtrFBL7 in secondary growth, the PtrFBL7 gene is transformed into poplar to generate the transgenic poplars overexpression and knock-out of PtrFBL7. The differences of phenotype and secondary growth among the transgenic and non-transgenic plants are analyzed. Yeast one hybrid and ChIP-PCR are employed for investigation of the upstream regulators of PtrFBL7. The genes effected by PtrFBL7 will be determined by the differentially expressed genes among transgenic and non-transgenic poplar plants using the RNA-Seq analysis. These results should be contributed in revealing the regulatory role of PtrFBL7 in secondary growth and provide the theoretical and practical base for molecular breeding of trees.

生长素在植物生长发育中通过信号转导途径来发挥重要的调控作用，而生长素受体是其信号转导的关键元件之一。我们已对杨树转运抑制响应蛋白（PtFBLs）基因进行了基因序列、系统进化关系以及相对表达模式等分析。发现PtrFBL7在维管组织和形成层中表达量较高，说明它可能在次生生长中起重要作用。为了揭示PtrFBL7在次生生长中的作用及调控机制，创建PtrFBL7基因过表达和敲除的转基因杨树，比较它们与非转基因杨树之间表型和次生生长的差异，确定其生物学功能；并通过酵母单杂交、ChIP-PCR技术等确定其上游的调控基因；同时，利用RNA-Seq技术比较转基因和非转基因杨树之间的基因表达差异，鉴定PtrFBL7所影响的下游靶基因。通过以上研究，阐明PtrFBL7基因在次生生长中的作用和调控机理，为林木材性改良育种提供依据。

木材形成是一个复杂的生物学过程，是维管形成层周期性活动的产物，是次生木质部连年沉积、附加的结果。在此过程中，植物激素起着重要的调节控制作用，如生长素、细胞分裂素和赤霉素等。其中生长素是一类具有极性运输特性的天然植物内源激素，主要通过信号转导途径来调控植物的生长发育，而生长素受体是其信号转导的关键元件之一。因此，开展生长素受体相关研究对于揭示生长素在次生生长中的作用机制具有重要意义。本项目以银腺杨（即84K杨，Populus alba × Populus glandulosa）为材料，克隆银腺杨PagFBL7（故将PtrFBL7更改为PagFBL7）基因，通过创制PagFBL7基因过表达、基因敲除等遗传材料，比较发现，2月龄过表达转基因植株的株高和茎部木质部宽度显著高于非转基因84K杨和RNAi抑制表达转基因植株，过表达转基因植株的成熟叶片明显向外卷曲，且叶绿素含量较高。利用酵母单杂交技术、瞬时表达技术等筛选、鉴定了其上游调控因子PagDofn，同时，利用RNA-Seq技术比较转基因和非转基因杨树之间的基因表达差异，鉴定PagFBL7所影响的下游靶基因，并发现PagFBL7能够与蛋白PagIAA3.3和PagIAA3.4相互作用。通过以上研究，阐明PagFBL7基因在次生生长中的作用和调控机理，为林木材性改良育种提供依据。