泡桐维管形成层活动周期相关microRNAs的鉴定与功能分析

The vascular cambium and its derivative secondary growth is a significant characteristic in woody plants. It is not the process of growth in the model plant Arabidopsis thaliana and rice. So it has important biological significance. Cambium activity periodicity plays a key role in the growth and development of the trees. However, so far it is not deep enough to understand the regulatory mechanism of cambial activity periodicity transition at the post transcriptional level. Our recent work demonstrated that miRNAs plays an important role in the regulation of growth and development of woody plant. On basis of our previous works, the miRNAs and its key target genes related to wood formation during cambium activity periodicity will be investigated in dormant, reactivating, and active cambium of Paulownia, respectively by using anatomy, molecular biology and bioinformatics methods. Furthermore, the role of key microRNAs in regulation of related to genes of development and differentiation in vascular cambium will be further explored, thus uncovering molecular mechanism of microRNAs regulation on cambium activity periodicity and laying the foundation for improving yield and quality of wood by using biological technology. The project is the first time to investigate the miRNAs and its key target genes related to cambium activity periodicity, which will help the further studies on the molecular mechanism of microRNAs regulations on the cambium activity periodicity in woody plants.

形成层及其衍生的次生生长是木本植物的显著特点，也是模式植物拟南芥、水稻中不具备的生长过程，因而具有重要的生物学意义。形成层的周期性活动在树木的生长发育过程中起着关键的作用，但目前对于调控形成层活动周期的分子机制在转录后水平上的了解还不够深入。我们近来的研究结果表明，miRNAs在调控木本植物的生长发育过程中起重要作用。本项目拟在前期研究基础上，分别以泡桐休眠期、萌动期和活跃期的形成层为材料，运用解剖学、分子生物学和生物信息学的手段系统研究和鉴定形成层活动周期相关miRNAs及其靶基因，解析重要miRNAs对形成层活动周期相关基因的调控作用，进而揭示miRNAs调控泡桐形成层周期性活动的分子机制，为利用生物技术手段改善木材产量和品质奠定基础。这是首次系统全面的分离和鉴定泡桐形成层活动周期相关miRNAs及其靶基因，对揭示木本植物形成层发育调控的分子机制研究具有重要的理论意义和潜在的应用价值。

维管形成层及其衍生的次生生长是木本植物的显著特点，也是模式植物拟南芥、水稻中不具备的生长过程，因而具有重要的生物学意义。泡桐是我国重要的优质速生用材树种。与拟南芥和水稻相比，泡桐的基因组非常庞大，但其维管形成层发育过程中miRNAs调控的分子机制还不清楚。本项目以休眠期、萌动期和活跃期的泡桐形成层为材料，运用解剖学、分子生物学和生物信息学的手段系统研究形成层发育过程中具有显著表达差异的miRNAs。研究结果表明，发现了20个保守miRNA家族和28个新的miRNA。利用RT-PCR和亚克隆测序的方法，验证了其中25个新的miRNA前体序列。荧光定量PCR结果揭示了10个保守和10个新miRNA在泡桐形成层不同发育阶段的特异表达，5' RACE的实验进一步证实了8个miRNA的靶基因。烟草叶片的瞬时表达实验表明，pto-miR171a和靶基因Unigene13248，pto-miR18和靶基因Unigene39201确实发生切割作用。这是首次系统全面的分离和鉴定泡桐维管形成层发育相关miRNAs及其靶基因，对揭示木本植物维管形成层发育调控的分子机制研究具有重要的理论意义和潜在的应用价值。