结合连锁分析和关联分析定位莲花期和地下茎QTL及候选基因鉴定

Lotus is an important ornamental aquatic plant. Flowering time is a primary determinant of the ornamental value of lotus,and is close related to the formation of rhizome. Studying the genetic mechanisms controlling flowering time and rhizome and revealing the genetic relatedness between them are important for guiding molecular breeding of new lotus cultivars with long flowering time. In this project, using SNP markers which is discovered by restriction site associated DNA sequencing (RAD-Seq), a genetic linkage map of lotus will be constructed based on an F2 population derived from two lotus cultivars with significant difference in flowering time and rhizome. According to the phenotype data of flowering time and rhizome from three years, QTLs controlling the two traits will be identified. The coincidental significant QTLs for flowering time and rhizome will be taken as the target QTLs. Markers and candidate genes which close linked with these two traits will be obtained through developing more markers in the confidence interval of the target QTLs. Associations of flowering time and rhizome with allelic loci in candidate genes will be tested by association mapping, and the function of candidate genes will be verified. The project will reveal the genetic variation of flowering time and rhizome formation, and lay a theoretical thesis for studying their gnentic mechanisms. The obtained molecular markers and functional genes which close linked with flowering time will provide useful information for molecular breeding of lotus cultivars with long flowering time.

莲是重要的水生观赏植物，花期长短是决定其观赏价值的重要指标。而花期与地下茎发育有密切关系，因此开展莲花期和地下茎发育遗传机制研究，解析两者之间的调控机理，对选育长花期莲品种具有重要的指导意义。本项目拟以已构建的亲本间花期和地下茎存在显著差异的F2分离群体为材料，利用RAD测序技术检测F2群体的SNP标记，构建莲花期遗传连锁图谱；完成对F2群体三年花期和地下茎表型性状的调查，检测相关QTL位点，选取同时控制花期和地下茎的QTL为目标QTL；通过目标QTL区间内分子标记加密及相关基因功能的分析，获得与目标性状紧密连锁的分子标记和候选基因；利用候选基因关联分析，检测花期和地下茎与候选基因的关联性，完成对候选基因的功能验证。项目将揭示莲花期和地下茎的遗传变异规律，阐明两者间的调控关系，为进一步解析莲花期遗传机制奠定基础；获得与花期紧密连锁的分子标记及基因，为长花期莲品种的分子育种提供理论依据。

莲是重要的水生观赏植物，花期长短是决定其观赏价值的重要指标。而花期持续长短与地下茎膨大启动早晚有密切关系。因此开展莲花期和地下茎发育遗传机制研究，解析两者之间的调控机理，对选育长花期莲品种具有重要的指导意义。通过本项目实施，获得结果如下：（1）连续3年调查F2分离群体的始花期、群体花期和地下茎性状等表型性状。各性状呈连续性分布，适合用于QTL定位研究；（2）利用全基因组重测序技术检测了F2分离群体的全基因组SNP标记，构建了包含1008个bin（22542个SNP）标记、8个遗传连锁群的莲高密度遗传连锁图谱；（3）结合表型性状与遗传连锁图谱，完成了始花期、群体花期和地下茎膨大系数的QTL 定位，得到22个显著性QTL。经整合，得到了6个年份间重复、贡献率高的目标QTL位点；（4）通过对目标QTL区间分子标记加密，结合莲基因组注释信息，筛选得到10个控制莲始花期、群体花期和地下茎膨大的候选基因；（5）开展了莲花期、地下茎的关联分析和比较转录组学研究，确定了莲开花和地下茎膨大的4个关键候选基因NnFT3、NnFT6、NnBLH6和NnGTE4；（6）完成了关键候选基因序列差异分析、时空表达模式、亚细胞定位研究，进一步通过转化拟南芥和马铃薯验证这些基因功能，发现超表达NnFT3可以导致拟南芥提前开花，超表达NnFT6延迟拟南芥开花，转化马铃薯验证NnFT、NnBLH6和NnGTE4功能正在表型鉴定和观察中。本项目获得与莲花期、地下茎膨大的关键候选基因，为长花期莲品种的分子育种提供理论依据，为进一步解析莲花期遗传机制奠定基础。