‘紫皮柚’果皮特异积累花青苷的调控机制

'Attractive, delicious and health-beneficial' are demanded features for citrus fruit breeding. Anthocyains contribute to the appearance of fruits by rendering them vivid red to purple. Moreover, as natural antioxidants, they are beneficial to human health. Our earlier germplasm survey has identified an unique ‘purple skinned pummelo’, which is the first case of pummelo accumulating anthocyanins in fruit peel, and one of the few citrus germplam which can synthesize anthocyanins worldwide. Distinct from blood orange, purple pummelo only accumulates anthocyanins in fruit peel. We identified the candidate regulatory gene CgMYB1, overexpression of this gene leading to massive accumulation of anthocyanins in Arabidopsis. In a further step, this project will investigate the regulatory mechanism of tissue specific accumulation of anthocyanins in fruit peel by characterization the interacting proteins and downstream target genes, elucidation the mechanism of response to light on the promoters, and develop purple-peel linked markers and apply to the fruit color breeding program. This study will elucidate the mechanisms of the tissue-specific regulation of anthocyanin in fruit. The outcome will provide the target gene and molecular markers for genetic improvement and breeding of novel appearance of citrus types.

培育‘好看、好吃、健康’的类型是柑橘育种的新需求。花青苷赋予果实艳丽的色彩，也是人类健康有益的天然抗氧化物。申请人前期发掘到‘紫皮柚’资源，为国际上已报道的第一例果皮积累花青苷的柚子资源（Citrus maxima），是柑橘属植物少有的积累花青苷的材料。与血橙不同，紫皮柚仅仅是果皮积累花青苷。项目前期鉴定了调控紫皮柚花青苷积累的候选基因CgMYB1，在拟南芥超量表达该基因导致花青苷大量积累。本项目拟进一步阐明紫皮柚果皮特异性积累花青苷的调控机制，筛选CgMYB1的互作蛋白及下游靶基因，明晰启动子对光信号途径的响应机制；开发紫皮性状紧密连锁的分子标记，用于筛选果皮富含花青苷的优良单株。本研究将进一步深化对柑果类果实花青苷组织特异性调控的认识，同时对于柑橘果皮色泽改良提高外观新颖性有应用价值。

项目前期鉴定了调控紫皮柚花青苷积累的候选基因CgMYB1，在拟南芥超量表达该基因导致花青苷大量积累。本项目拟进一步阐明紫皮柚果皮特异性积累花青苷的调控机制，筛选CgMYB1的互作蛋白及下游靶基因，明晰启动子对光信号途径的响应机制；开发紫皮性状紧密连锁的分子标记，用于筛选果皮富含花青苷的优良单株。本研究将进一步深化对柑果类果实花青苷组织特异性调控的认识，同时对于柑橘果皮色泽改良提高外观新颖性有应用价值。主要研究结果如下：.1)组装了紫皮柚高质量参考基因组；通过套袋实验，发现光照能诱导血橙和紫皮柚果皮花青苷积累。血橙CsRuby1启动子是一个既受光照诱导，也能受低温诱导，而紫皮柚CgRuby1启动子只受光照诱导；.2)定位了CgRuby启动子光响应关键区域，结果表明CgRuby1启动子ATG上游的0bp到-234 bp区域是响应光照诱导的关键区域；.3)提出了血橙和紫皮柚花青苷合成受光照和低温诱导的调控模型。通过对不同柑橘的Ruby1启动子进行序列分析，鉴定到启动子上均含有保守的G-box光响应元件。此外，凝胶电泳迁移和双荧光素酶实验结果显示HY5可以结合G-box元件，从而正向调控Ruby1的表达来促进花青苷的积累；.4)利用紫皮柚、血橙作为杂交亲本创制遗传群体，开发分子标记辅助苗期选择紫色性状单株；筛选出“琯溪蜜柚×紫皮柚”杂交群体紫皮红肉优良单株。