Ttu-miR9772调控野生二粒小麦耐盐性的分子机制研究

miRNA作为一类重要的调控因子，在植物逆境响应与耐受过程中发挥重要作用。野生二粒小麦是小麦最重要的种质资源和基因库之一。发掘、利用其优异耐盐miRNA对丰富小麦耐盐基因资源、克服遗传同质化具有重要意义。本研究以前期发现的野生二粒小麦耐盐关键miRNA Ttu-miR9772为研究对象，首先克隆其前体和启动子序列，并利用生信预测和剪切产物5’RACE鉴定其准确的靶基因及剪切位点，结合Ttu-miR9772及其靶基因的表达特性分析，初步阐明其产生和表达的调控机制；在此基础上，利用扩增子测序技术分析其前体、启动子及靶基因在不同耐盐性材料间的遗传变异，结合耐盐性关联分析，鉴定关键SNP位点及优异等位变异；最后，利用过表达和短串联靶标模拟(STTM)技术，对其生物学功能进行系统研究，以期阐明Ttu-miR9772调控野生二粒小麦耐盐性的分子机制，为利用小RNA改良小麦及其他作物的耐盐性服务。

As an important regulator, miRNA plays an indispensable role in regulating stress response and tolerance in plants. Wild emmer wheat (Triticum dicoccoides Korn) is one of the most important resources and gene pools for wheat genetic improvement. It is of great significance to exploit the salt tolerance responsive miRNAs from wild emmer wheat to enrich genetic resources and overcome the genetic homogeneity for developing salt tolerant varieties in wheat. In this study, the Ttu-miR9772 is selected to investigate its function and mechanism in regulating salt tolerance in wild emmer wheat, which is one of key salt-responsive miRNAs as found by our previous study. Firstly, the precursor and promoter sequence of Ttu-miR9772 is cloned, and its target gene and splice sites are obtained by bioinformatics prediction combined with 5'RACE approach. Then, the expression profiles of Ttu-miR9772 and its targets are systematically investigated. Based on these results, the processes of its production, expression and regulation is preliminarily clarified. Furthermore, the genetic variations of the precursor, promoter and target of Ttu-miR9772 are revealed by amplicon sequencing, and then are associated with salt tolerance to identify the salt tolerance related loci and excellent alleles. Finally, the biological functions of Ttu-miR9772 is revealed using over-expression and STTM (short tandem targets mimic) methods. This study aims to elucidate the molecular mechanism of Ttu-miR9772 regulating salt tolerance in wild emmer wheat, which will contribute to improve the salt tolerance of wheat and other crops through making use of small RNAs.