基于高密度遗传图谱构建的金叶弯刺蔷薇叶色变异相关候选基因定位与挖掘

The leaf-color mutation of plants frequently occurs in the nature, which provides the most important opportunity to obtain the colorful-leaf ornamentals. However, few studies had been focused on the molecular mechanism of leaf-color mutation among the ornamental plants, especially the woody ornamentals. The yellow-leaf (yl) mutant of Rosa beggeriana is a bud sport of wild rose species R. beggeriana by the treatment of γ-ray irradiation and the genetic analysis indicated the mutated trait should be dominant & monogenic inheritance. In this project, a good example could be established for the analysis of molecular mechanism of leaf-color mutation among the ornamentals through the mapping and cloning of the gene involved in the yl mutant of R. beggeriana. Firstly, the high-throughput molecular markers will be explored for the construction of high-resolution of genetic map of Rosa beggeriana yellow-leaf mutant × Rosa davurica based on the completed rose genome and the re-sequencing data of the parents. The traits associated with leaf color, flower color and inflorescence could be located on the genetic map and the rose genome. Furthermore, the molecular markers closely related to the yellow-leaf trait will be further explored for the fine mapping of the yl gene. Consequently, the candidate genes could be predicted in the light of the rose genome and the gene expression differences based on the RNAseq of Rosa beggeriana and its yl mutant. The candidate genes will finally be cloned and the gene function would be further identified with the transformation systems of Arabidopsis and rose.

自然界广泛存在的叶色变异现象是获得彩叶观赏植物新种质的重要来源。但是，目前有关观赏植物尤其是木本观赏植物的叶色变异分子机制的深入研究较少。金叶弯刺蔷薇是新疆弯刺蔷薇通过辐射诱变获得的叶色突变材料，遗传分析表明该突变性状由显性单基因遗传控制。以金叶弯刺蔷薇为研究对象，通过图位克隆的方法进行叶色突变性状相关基因的定位与挖掘，对于深入阐明其叶色变异分子机制，以及新型观叶月季品种选育具有重要意义。本项目将围绕这一特色变异种质，充分利用月季基因组测序计划获得的全基因组数据以及亲本重测序数据，开发高通量分子标记进行金叶弯刺蔷薇×山刺玫高密度遗传图谱构建，结合表型观测数据进行目标性状相关基因的初步定位；在此基础上继续开发分子标记加密初定位区域，完成金叶突变性状相关基因的精细定位，结合月季全基因组及金叶弯刺蔷薇转录组测序信息，预测控制突变性状的候选基因，并开展基因克隆和功能验证分析。

自然界广泛存在的叶色变异现象是获得彩叶观赏植物新种质的重要来源。但是，目前有关观赏植物尤其是木本观赏植物的叶色变异分子机制的深入研究较少。金叶弯刺蔷薇是新疆弯刺蔷薇通过辐射诱变获得的叶色突变材料，遗传分析表明该突变性状由显性单基因遗传控制。本项目首先通过表型、生理指标测定以及转录组测序等方法，明确了金叶弯刺蔷薇叶色变异的形态解剖、生理生化与分子基础。以金叶弯刺蔷薇×山刺玫F1杂交群体为材料，通过BSAseq混池测序结合MassArray技术进行SNP分型，利用36个有效SNP标记和1个叶色突变标记完成总图距为34.4cM的遗传连锁图谱构建，其中金叶突变性状定位于上述遗传图谱的22.7cM处，分别距离上、下游标记为0.2cM和0.4cM；对照参考基因组物理图谱，将该性状定位在中国月季‘月月粉’（R. chinensis ‘Old Blush’）基因组CM009586.1号染色体的6,290,023bp-6,581,773bp之间，区间总长度为0.28Mb，该区域内含有非同义突变SNP位点的基因14个，移码突变InDel位点的基因5个。通过对候选基因克隆与表达模式分析，以及初步的功能验证分析，我们认为转录因子RbERF1可能为关键候选基因。本项目的科学意义主要在于金叶弯刺蔷薇作为蔷薇属植物中挖掘创制的第一个叶色变异材料，利用经典的正向遗传学，即基于遗传图谱构建的图位克隆技术，开展金叶弯刺蔷薇叶色变异相关候选基因的精细定位及功能验证分析，对于阐明金叶弯刺蔷薇叶色变异的分子机制，培育新型观叶月季品种具有很重要意义。