转录因子MYB75调控矢车菊素糖苷累积介导文冠果花色转变的分子机制

The flower color transformation has important value on breeding and ornamental, but the molecular regulation mechanism is not clear because of the flower color model plants do not have the transformation characteristics. Xanthoceras Sorbifolium is fine tree with flower and fruit using in the north of China, the change of flower color is obvious and the period from seedlings to flowering is short. It is an ideal material for studying the regulation of flower color on flower trees.In the early stage of the project, the self-cultivated new red flower varieties were used as test materials. In the early stage of this project , we have performed an integrated analysis of the transcriptome and the key components of the base petal anthocyanins in different periods. Results shown that the accumulation of cyanidin lucosidein is a key factor in the transformation of flower color, which is closely related to the transcription factor MYB75. To verify the functionality of XsMYB75 in the regulation of anthocyanin biosynthesis，over-expression and CRISPR/Cas9 vectors will transient expressed and identified. Meanwhile, we attempt to generate transgenic lines expressing over-expression , CRISPR/Cas9 and wild-type respectively, and anthocyanins will be tested by HPLC-MS .At the same time, the petal samples of multiple times from expressing plants were collected for transcriptome sequencing, downstream regulatory genes were screened, and downstream direct target genes were determined by Chromatin immunoprecipitation and Electrophoretic mobility shift assay. Further elucidating the molecular mechanism of XsMYB75 regulating flower color transformation, which can provides a theoretical basis for the creation of new ornamental cultivar in Xanthoceras sorbifolium.

花色转变具有重要的繁育意义和观赏价值，但由于模式植物花色不具备转变特性，相应的分子调控机制尚不明确。文冠果是我国北方优良的花果兼用树种，花色转变明显，幼苗达到开花周期短，是林木上研究花色调控的理想材料。项目前期以自主培育的内红型大花新品种为试材，进行4个时期基部花瓣花青素关键组分和转录组测序分析，证实文冠果花色转变的关键因素是矢车菊素糖苷的有效累积，并与转录因子MYB75密切相关。本项目拟通过XsMYB75基因克隆，构建过表达载体和CRISPR/Cas9载体，获得文冠果遗传转化植株，进行表型观察、矢车菊素糖苷含量测定；同时采集过表达植株多时期花瓣样品进行转录组测序，筛选和鉴定下游调控基因，结合染色质免疫共沉淀和凝胶迁移等方法确定下游直接靶基因，进而阐明XsMYB75调控花色转变的分子机制，为观赏型文冠果新品种的创制提供理论依据。

花色转变具有重要的观赏价值和繁育意义，但由于模式植物花色单一不具备转变特性，相应的分子调控机制尚不明确。文冠果是我国北方优良的花果兼用树种，“白花型”和“红花型”花瓣基部花色逐渐变红，而且两者花色着色范围存在显著差异。项目通过全面检测“白花型”文冠果多时期花青素和类胡萝卜素含量，显示矢车菊素-葡萄糖苷和芍药花素-葡萄糖苷是"白花型"花色变红的关键因素，联合基因表达数据关联分析发现XsMYB58、XsGST、XsANS和XsFLS与上述代谢物存在显著相关性。同时项目还基于已有的重测序数据，解析了“白花型”和“红花型”文冠果形成的遗传学基础，全基因组关联分析显示花色类型紧密相关的突变位点主要和9-顺式环氧类胡萝卜素双加氧酶、谷胱甘肽转移酶、谷氨酸受体、黄烷酮、查尔酮黄烷酮异构酶、油菜素内酯转录组因子相关，这些基因位点突变在花色性状分化中起到重要作用，从而形成着色范围显著差异的两种花色类型。花瓣颜色转变和花色类型形成机制为观赏型文冠果新品种的创制提供参考。