内源诱捕靶标介导MIR171调控百合体细胞胚发生的功能解析

体细胞胚发生对于百合种球繁育、种质保存、遗传改良等均具有重要意义。miRNA作为基因上层调控因子在体胚发生中起重要作用，而这种调控受内源诱捕靶标（eTM）的介导，但eTM介导miRNA调控植物体胚发生的机制尚不明确。本研究在确定miR171参与调控百合体胚发生并初步验证其对靶基因SCL6的调控作用、预测获得介导因子lpu-eTM171的基础上，拟通过茎环反转录、RACE及qRT-PCR技术克隆miR171成熟体及前体、eTM和靶基因，并分析其差异表达模式；通过启动子顺式作用元件功能分析明确miR171转录特性；通过功能获得性和缺失性研究，分析过表达、沉默eTM和miR171的转基因百合中miR171、SCL6的差异表达水平；注释三者相关功能并进行特异代谢调控通路分析，阐明eTM-miR171-SCL6参与调控百合体胚发生的机理，为明确百合体细胞胚发生机制、并实现体胚发生的人工调控奠定基础。

Somatic embryogenesis is vital to the bulb propagation, germplasm preservation as well as genetic traits improvement in Lilium. miRNA is important in the process of somatic embryogenesis as the gene’s upper control factor. And the function of miRNA on target genes is regulated by endogenous target mimics (eTM). Until now the mechanism of eTM mediating somatic embryogenesis in plants is not yet clear. Our research group has identified miR171 involved in somatic embryogenesis on Lilium, and the target genes SCL6 as well as lpu-eTM171 have also been verified and predicted successfully in our recent work. In this project, we intend to carry out the following work. Cloning the full length cDNA and analyzing the expression profiles of miR171, pre-miR171, eTM, and SCL6 by stem-loop RT-PCR, RACE and qRT-PCR; Explicating the transcription characteristics of miR171 via the functional analysis of the promoter cis-function element; Applying the gain-of-function and loss-of-function study to construct the express profiles of miR171 and SCL6 in over-expression and RNAi transgenic plants of eTM and miR171; Annotating the function and analyzing the specific metabolic regulation pathways of miR171, eTM and SCL6, to clarify the regulating mechanism of eTM-miR171-SCL6 during somatic embryogenesis in Lilium. This study may establish foundation for making clear the mechanism of somatic embryogenesis and its artificial regulation.