红花草莓MYB类转录因子调控花瓣花青素苷合成的分子机理

The flower of the genus Fragaria is white. The red-flowered strawberry, a new kind of ornamental and edible strawberry, is an intergeneric hybrid from the cross of F. × ananassa and Potentilla palustris. It is of great ornamental value with red flowers, red fruits and green leaves. In our previous study, cyanidin 3-glucoside was regarded as the main anthocyandins in the petals of red-flowered strawberry. Four MYB transcription factors might also be responsible for anthocyandin synthesis regulation in the petals of red-flowered strawberry based on our previous completed RNA-seq sequencing of red-flowered petals and white-flowered petals in hybrids from the cross of Pink Princess × Pink Beauty. In this study, full-length gene of four selected MYB transcription factors will be isolated. To analyze the correlation between the expression of MYB and anthocyadin levels in the petals, the temporal and spatial regulation expression patterns of MYB transcription factors and anthocyandin contents will also be investigated using real-time PCR and HPLC-MS, respectively. And subcellular localization will be used to analyze their nuclear localization characteristics. The function of MYB transcription factors in the regulation of anthocyandin synthesis will be further tested by transient expression and genetic transformation of tobaccos, and gene silencing by VIGS in the petals of red-flowered strawberry cv. ‘Sijihong’. The binding specificity of MYB transcription factors to cis-acting element and their transcriptional activities will be investigated by using electrophoretic mobility shift assay and a yeast one-hybrid system to clarify the MYB regulation molecular mechanism of anthocyandin synthesis in red-flowered strawberry. The results will be not only to provide theoretical support for breeding new red-flowered strawberry cultivars with rich anthocyadins, but also to help to enrich regulation theory of anthocyadin biosynthesis

草莓属植物均开白花，红花草莓是一种观赏兼食用的新型草莓，为属间杂种（Fragaria × Potentilla），其花红、果红、叶绿，具有很高的观赏价值。本项目前期明确了红花草莓花瓣的主要呈色物质是矢车菊素-3-葡萄糖苷，并基于红花草莓杂交后代红色和白色花瓣的转录组测序，筛选了4个调控花青素苷合成的MYB家族成员。在此基础上，本研究将分离这4个MYB转录因子的基因全长序列，利用实时荧光定量和亚细胞定位分析MYB时空表达模式与花青素苷合成的关系及其核定位特性；利用烟草瞬时表达和遗传转化、VIGS技术沉默红花草莓花瓣MYB基因，确定MYB家族成员的功能；利用EMSA凝胶阻滞和酵母单杂交分析鉴定启动子区域的关键顺式元件和MYB转录激活活性，揭示其调控花瓣花青素苷合成的分子机理。研究结果不仅对于培育富含花青素苷的红花草莓品种提供理论支撑，还有助于丰富植物花青素苷生物合成与调控理论。

红花草莓是一种新型的观赏草莓，为属间杂种（Fragaria × Potentilla），可观赏兼食用，其红花性状是从开红花的沼委陵菜（Potentilla palustris）通过远缘杂交导入到开白花的栽培草莓（Fragaria × ananassa）中去的。本项目以红花草莓品种‘四季红’为试验材料，通过对红花草莓杂交后代红花、白花的转录组测序和八倍体草莓MYB基因全基因组分析，筛选到了4个与红花草莓花瓣花青素形成相关的MYB家族成员，分别为FpMYB1、FpMYB86、FpMYB90、FpMYB308，发现这4个转录因子均定位在细胞核，且FpMYB90和FpMYB308基因在花瓣中特异表达，蕾期的表达量最高；通过构建载体，对烟草和红花草莓花瓣进行瞬时侵染，发现FpMYB90基因可以促进花青素的积累；分析了红花草莓花蕾发育不同阶段miRNA差异表达情况，筛选得到了33个与红花草莓花色形成相关的候选miRNAs和其对应的40个靶基因。这些研究为深入解析红花草莓花瓣着色的调控机理奠定了理论基础。本项目完成了预期任务，取得了较好的科研成果，发表论文5篇，其中4篇为SCI；获得国家发明专利1项；培养博士研究生1名、硕士研究生3名。