苹果高类黄酮优异种质创制及其类黄酮代谢机理的研究

The creation of excellent apple germplasm with high flavonoid and easy coloring and the breeding of characteristic varieties of high-flavonoid apples are important technical need for apple industry. This application is based on the creation of high flavonoid germplasm ‘CSR6R6’, the construction of hybrid population and the obtainment of candidate genes such as HsfA8, TMT1/2 and ARF3 by phenotypic analysis, RNA-seq and gene expression analysis. Further through the primary selection, the second selection and the final selection procedures, new functional apple cultivars were bred. The germplasm of CSR6R6 and its hybrid progeny lines were took as materials. We studied the structural characteristics and temporal and spatial expression of candidate genes and their relationship with the content of flavonoids in fruits; Yeast two hybrids, EMSA and other techniques will be used to study the protein-protein and protein-DNA interaction characters and regulation functions between candidate genes and structure genes, and transgenic function verification will be performed. The relationship between heat shock transcription factor, glucose transporter, auxin signaling factor and flavonoid metabolism was studied in high-flavonoid apples to investigate their flavonoid metabolism mechanism, so as to provide basic data for the enriching apple resources and the theoretical system of flavonoids metabolism and provide the scientific basis for the breeding of high-flavonoid apples.

创制高类黄酮优异种质、培育易着色的高类黄酮苹果等特色多样化品种是苹果产业的重要技术需求。本申请在高类黄酮优异种质CSR6R6创制及其杂种后代分离群体构建以及通过RNA-seq与基因表达分析获得HsfA8和TMT1/2等候选基因的基础上，进一步通过初选、复选和决选程序，选育高类黄酮苹果新品种；以CSR6R6及其高类黄酮苹果新品系为试材，通过基因克隆和生物信息学分析等研究候选基因结构特征和时空表达特性及其与参试品种果实类黄酮含量变化的关系；利用酵母双杂交及EMSA等技术研究候选基因与FLS等相关基因在蛋白-蛋白和蛋白-DNA水平上的互作特点及调控作用，并进行转基因功能验证，研究热激转录因子、糖转运蛋白和激素信号因子与高类黄酮苹果类黄酮代谢的关系和作用机理，从而探讨高类黄酮苹果优异种质的类黄酮代谢机理，为丰富苹果资源及苹果类黄酮代谢机理的理论体系提供基本资料，并为高类黄酮苹果育种提供科学依据。

创制高类黄酮优异种质、培育易着色的高类黄酮苹果等特色多样化品种是苹果产业的重要技术需求。本申请在高类黄酮优异种质CSR6R6创制及其杂种后代分离群体构建以及通过RNA-seq与基因表达分析获得HsfA8和TMT1/2等候选基因的基础上，进一步探究了苹果类黄酮的代谢机理，并选育出系列高类黄酮苹果新品种。课题所有研究内容均按计划进行，主要取得了如下研究成果：①以高类黄酮优异种质CSR6R6与‘红富士’和‘嘎啦’等苹果品种杂种后代分离群体为研究对象，对前期初选的株系进行嫁接，建立高接鉴定圃，通过对各参试株系果实大小与形状、糖、酸、香味物质、类黄酮和矿物质含量及硬、脆度等品质因素的检测与评价，并通过初选、复选和决选程序，最终培育出果肉全红、类黄酮含量高、易着色、营养保健价值大、鲜食品质优良或加工性能优异的‘幸红’‘福红’‘美红’‘满红’等高类黄酮苹果新品种4个，均通过山东省良种审定，获得植物新品种权6项；②以CSR6R6及其高类黄酮苹果新品系为试材，探究了MdHsfA8a/MdHsf3b/MdHsf4a等热激转录因子、MdTMT1等糖转运蛋白、MdARF13/MdERF1B等激素信号因子以及MdMYBPA1/MdMYB6等MYB转录因子与高类黄酮苹果类黄酮代谢的关系和作用机理。系统解析了热激转录因子、糖转运蛋白和激素信号因子与高类黄酮苹果类黄酮代谢的关系和作用机理，为进一步丰富苹果资源、完善苹果类黄酮代谢机理的理论体系和调控网络提供基本资料，并为高类黄酮苹果育种提供科学依据。③课题共发表项目资助号的论文46篇，其中SCI论文35篇（其中一区9篇，影响因子>10的1篇，影响因子>5的22篇），授权发明专利4项（国际专利1项，国家发明专利3项），获植物新品种权6个，审定高类黄酮苹果新品种4个，推广及应用前景广阔；项目执行期间培养博士后1人，培养博士11人，其中毕业7人，在读4人，培养硕士10人，其中毕业9人，均超额完成项目计划任务指标。课题相关研究成果作为核心内容荣获2020年度国家技术发明二等奖1项。