四倍体月季高密度遗传连锁图谱构建及花色基因的分子标记定位

Roses (Rosa spp.), which belong to the family Rosaceae, are one of the most important ornamental flowers in terms of economy and cultural history of humankind, due to their beauty, attractiveness and different usage areas. Besides, flower color is one of the important quality characteristic traits of ornamental plants that has been paid close attention. Compared with traditional breeding method that is regarded as higher degree of uncertainly of unpredictability, molecular marker assisted selection is supposed to be a more efficient way to enhance and accelerate the breeding process. Genetic maps provide a framework of the chromosomal organization and can be used to identify genetic linkage between markers and desirable traits, to locate single genes, for QTL analyses and serve as starting point for map-based cloning of genes etc. Currently, related research are mainly focused on diploid level, while in this genus polyploidy occurs frequently in wild as well as in cultivated roses. The majority of the wild species are diploid, whereas most cultivated roses are tetraploid or triploid. QTLs are difficult to select for in a highly heterozygous tetraploid genome like that of the cultivated rose. Thus, this research aims to 1) construct a high-density genetic linkage map in tetraploid level, 2) map specific QTLs that are related to flower color and detect the candidate genes. Definitely, a better understanding of a genetic linkage map with a tetraploid population and important genes that contribute to desired phenotypes will lead to improved rose cultivars and provide theoretical basis for map-based gene cloning.

月季是蔷薇科蔷薇属的一个重要类群。因其姿态优美，色彩绚丽，用途众多等优点，被公认为重要的观赏植物之一。而观赏植物的花色又是其中一个重要品质特征，被广泛关注。传统的育种工作存在很大的盲目性和不可预测性，而分子育种可以显著提高育种效率，加速育种进程。遗传连锁图谱的构建可为性状定位、分子标记辅助选择育种、功能基因克隆等提供技术平台，具有不可替代的作用。目前，蔷薇属中图谱的构建多集中在二倍体水平。然而在本属植物中，野生原种以及栽培品种的多倍体现象频发，现代月季品种绝大多数为四倍体。当在前人所构建的二倍体图谱上进行性状定位时，效率往往很低，品种改良进程依旧缓慢。因此，本项目拟通过（1）构建四倍体水平上高密度遗传连锁图谱，（2）并在此基础上定位影响花色性状的QTLs并挖掘与其相关的候选基因。研究结果旨在为月季分子标记辅助育种和花色基因工程改良提供科学依据，为今后开展基因图位克隆奠定理论基础。

以月季‘太阳城’和‘云蒸霞蔚’杂交获得的F1群体以及不同颜色现代月季品种为材料，利用HPLC-PDA-MS方法，建立月季花瓣中类黄酮色素和类胡萝卜的快速提取、分离和鉴定的技术体系并对月季F1作图群体花瓣中的色素进行定性定量分析；利用简化基因组测序技术手段对全部子代进行测序，共得到125.92 Gb原始数据，利用GATK进行变异检测，筛选父母及子代大于10×的标记，获得17,382个SNP标记。加上前期课题组开发的381个SSR和AFLP标记，去除没有分离的标记，最终获得6,885个高质量的标记，以此来构建月季高密度遗传图谱，包含7个连锁群，总图距为1,158.90 cM，标记有6,842个，标记间平均遗传距离为0.18 cM。此外，结合测定的表型数据，利用Polyqtl进行QTL分析，共检测到50个稳定的与色差紧密关联的QTL位点，与花青苷含量相关的有40个，与黄酮醇含量相关的有113个，与类胡萝卜素相关的有79个，共检测到11个QTL簇与类黄酮代谢紧密连锁，6个QTL簇与类胡萝卜素代谢紧密连锁，进一步筛选出12个高花青苷含量位点，6个高类胡萝卜素位点。最终筛选出41个类黄酮代谢候选基因，20个类胡萝卜素代谢候选基因。