调控柚果实类胡萝卜素代谢的关键基因挖掘

Fruit color is important for both fruit appearance and nutrition trait. Citrus fruits are rich in carotenoids, and present various colors since with different types and contents of carotenoids. The fruit color of normal pummelo is light yellow or white. The red-, yellow-fleshed and red-albedo mutants found and used in the industry were unusual fruit traits in citrus and attracted broad attentions. These mutants are of significance for their economic, nutritional and research values, particularly ideal materials for the investigation of regulatory mechanism of carotenoid metabolism and fruit color formation. The fruit color of pummelos show abundant variations, but so far the knowledge on the genes in essence regulating carotenoid metabolism is limited. This research will focus on the identification of key genes in controlling carotenoid metabolism, with a unique set of color mutants of pummelo (including yellow-, red-fleshed and red-peeled mutations) recently found in China as the materials. Research contents will include three aspects i.e. the genetic loci responsible for the carotenoid accumulation, transcriptional regulatory factors of carotenoid biosynthetic genes, and the plastid development & differentiated genes during the storage of carotenoids. The candidate genes will be confirmed by functional transformation in Kongkong kumquat (it can set fruits within one year after seed germination), and model fruit crop tomato. Their effects on the regulatory mode and potential in genetic improvement will be evaluated. This research is to transform the unique germplasms into effectual gene(s); at the same time, the results will provide new insight into the molecular mechanism of fruit color mutation, and strengthen the base for the genetic improvement of fruit color.

果实色泽既是外观性状又是营养性状。柑橘果实富含类胡萝卜素，因含有的种类和量的不同呈现各种色泽。柑橘中的柚类，其果肉一般为乳白或淡黄色，近年生产中发现并应用的红肉、黄肉和红皮体细胞突变体是研究果实色泽性状形成与调控机制的良好材料。柚果实色泽变异丰富，但目前对变异原因即类胡萝卜素代谢的关键调控基因还不清楚。本项目拟利用我国特有的一套柚不同色泽（红肉、黄肉和红皮）突变体为材料，从控制色泽变异的遗传位点、调控类胡萝卜素合成基因的转录因子，以及控制类胡萝卜素贮存亚细胞器（有色体）发育的关键基因等三方面开展研究，旨在解析控制柑橘类胡萝卜素代谢的关键因子，发掘关键基因；通过转化播种当年能开花结果的山金柑以及模式作物番茄，评价基因对类胡萝卜素代谢的调控模式、作用及遗传改良潜力。本研究是将我国特有的果树遗传资源转换为基因资源；同时，对深入理解果实色泽变异的分子机制，以及指导色泽遗传改良均具有重要意义。

项目执行5年来，项目组利用色泽突变体、杂交群体和自然群体系统研究了柑橘类胡萝卜素代谢调控机制，取得了系列重要成果。分别利用RAD-Seq技术和SLAF标签简化基因组测序构建了黄肉琯溪蜜柚×坪山柚和红橘×枳杂交群体的高密度遗传图谱；利用BSA-Seq、GWAS、转录组、代谢组等手段挖掘柑橘果实色泽主效位点，从中筛选出6个果肉色泽性状候选基因，鉴定了红色果皮特有的C30 脱辅基类胡萝卜素——β-柠乌素合成的关键基因CCD4b；解析了类胡萝卜素生物合成关键酶基因LCYB的转录调控机制，挖掘到直接调控类胡萝卜素合成的转录因子CsMADS6（Plant Physiology, 2018）和CsMYB68（New Phytologist, 2017）；首次提出柑橘有色体分化模型（Plant Physiology, 2015），解析了柑橘果实类胡萝卜素积累的亚细胞基础，获得有色体分化和贮存的候选基因CitABC1K、 FBN1和Or基因；开发了分子标记用于果实色泽育种。在Plant Physiology、New Phytologist、DNA research等刊物发表SCI论文18篇，其中IF>5的论文6篇；授权发明专利1项、新品种权2项、软件著作权1项，审定新品种1个，以本项目部分研究成果作为核心内容的科技成果“柑橘基因组及果实色泽品质形成机制”获教育部自然科学一等奖（2018），项目主持人获2017年度湖北省科学技术突出贡献奖；关于柑橘色泽改良的研究论文被评为第二届中国科协优秀科技论文；培养博士研究生9人，硕士研究生4人。项目圆满完成了研究任务，所获得的研究结果在理论上丰富了果实类胡萝卜素代谢的调控与存贮机制，为未来色泽分子设计育种提供了基因资源和理论基础。