miR159介导SbMYB1调控黄芩苷合成代谢的分子机制研究

Baicalin is one of the main active ingredients of Scutellaria baicalensis with wide pharmacological functions, but its exploration has been greatly limited due to its low yield. It is generally recognized that genetic modification of the biosynthesis pathway is an ideal strategy for high product of medicinal herbs resource. Our research group has cloned several key enzyme genes involving baicalin biosynthesis pathway and verified their roles in regulating secondary metabolism; meanwhile we found that miR159 effectively induced the expression of MYB1 transcription factor which as a result influenced the accumulation of flavonoid compounds. Based on the previous results, in this study, we plan to clone microRNA159 and its target gene MYB1, study their molecular functions as well as investigating their expression levels in different tissue of different developing period, then introduce miR159 and MYB1 individually and together into S. baicalensis by agrobacterium tumefaciens- mediated transformation method, detect the genes and proteins expression level of key enzymes involving flavonoid biosynthesis as well as measuring the accumulation level of baicalin, so as to explain the effect of miR159 and its target gene MYB1 on baicalin anabolism and the underlying regulating mechanisms. As a result, this study could provide new candidate targets and theoretic basis for metabolism engineering of cultivating new medicinal species with high baicalin yields.

黄芩苷是黄芩（Scutellaria baicalensis）的主要活性成分，具有重要药理功效，但其含量较低限制其广泛应用，而通过遗传改造其生物合成途径的方法被认为是获得高产药源的理想策略。本课题组克隆和鉴定了黄芩苷生物合成途径中多个关键酶基因，并初步验证了其次生代谢调控功能；同时初步发现植物miR159能有效调节MYB1转录因子的表达，且影响黄酮化合物的积累。故本项目拟进一步克隆黄芩microRNA159及其靶基因MYB1，明确其分子功能及发育时期和组织差异的表达模式，并基于根癌农杆菌介导的遗传转化方法单独/协同调控miR159和MYB1的表达水平，考察黄芩苷合成途径中关键酶基因的mRNA表达谱和蛋白表达谱及终产物积累情况，揭示miR159介导MYB1调控黄芩苷合成代谢的作用和分子机制，从而为培育高产黄芩苷新型药源的代谢工程提供候选靶点和理论基础。

黄芩苷作为一种源于传统中药的天然产物，具有广泛的药理作用，包括抗氧化、抗肿瘤、消炎抑菌、免疫调控、降压清脂、利尿利胆和保护心脑血管等, 其开发前景十分广阔。本研究利用分子生物学方法遗传改造黄芩苷代谢途径，并在黄芩苷生物活性和药物特性等方面进行了深入扩展，优化了项目结构和推进了项目意义。完成miRNA159和SbMYB基因的分离和载体构建，完成基因的遗传转化研究，分析miRNA159和SbMYB在黄芩中的信号调控网络，对黄芩苷的生物合成和代谢通路进行了深入解析，并从现代中药资源学的角度将生物学、药学和医学等多学科紧密结合，从生物信息学、细胞工程、基因工程、种质改良、品质评价和药效药理等全链条一体化设计，形成以黄芩苷为核心的资源开发和大健康产业研究体系。