基于表观遗传调控因子miRNA解析冬凌草道地性形成分子调控机制

冬凌草道地性明显，不同产地冬凌草中特征性功效成分冬凌草甲素（Oridonin，Ord）含量差异显著，是其道地质量属性的主要体现，但调控机制尚不明确。研究表明，冬凌草IrKSL基因的可变剪切引起其二萜类成分变化，而表观遗传调控因子miRNA通过调控靶基因表达参与植物二萜生物合成。推测冬凌草miRNA通过响应环境调控酶基因差异表达，引起不同产区冬凌草二萜类成分变化，是其道地性形成的重要机制。本研究在转录组基础上，结合sRNA和降解组测序，鉴定冬凌草中miRNA及其靶标，采用qRT-PCR、RLM-5' RACE、烟草瞬时转化技术以及拟南芥和冬凌草过表达miRNA，阐明冬凌草miRNA与靶基因的交互作用及miRNA生物学功能；构建miRNA介导的网络调控模型“miRNA-mRNA-Ord”，为解析冬凌草道地性形成机制提供新的思路，也为其他道地药材的机制研究提供参考。

The content of oridonin in Isodon rubescens from different habitats are significantly different, which basically reflect the essences of geo-herbalism, but the formation mechanism is not clear. A recent studies shows that alternatively spliced transcripts of IrKSL generate different types of diterpenoid compound. Epigenetic regulator miRNA can regulate diterpenoids by controlling the expression of different splice variant from the same target. It is speculated that miRNA controls the terpene synthase gene transcripts differentially expressed respond to environment change，thus result in different structure and content of diterpenoids, which may be the important mechanisms of geo-herbalism quality formation in Isodon rubescens. On the basis of transcriptome analysis, miRNAs and targets in Isodon rubescens leaves will be identification by small RNA sequencing and degradome sequencing. The cleavage and interaction of candidate key miRNA with its target will be illuminated by qRT-PCR, RLM-5' RACE and agrobacterium-mediated transient co-transformation assay, and the biological function of miRNA will be verified by cloning the precursor gene of miRNA and transformed it into Arabidopsis thaliana and leaf of Isodon rubescens by agrobacterium-mediated. MiRNA mediated “miRNA- mRNA- diterpenoids” regulatory network will be established to expound geo-herbalism forming mechanization of Isodon rubescens. The research will provide a new perspective for geo-herbalism forming mechanization study of other Chinese herbal medicine.