基于高密度整合图谱构建的石斛比较基因组学研究

Dendrobium is one of the rare traditional Chinese medicinal materials, the market demand is large, however, the current situation of the shortage of natural resources requires we take measures to actively respond to. Based on the fundamental research situation and our research basis of Dendrobium, the suggested project aims at going from the integration genetic linkage maps construction, we carry out the comparative genomic study of Dendrobium. Considering the premise of Dendrobium genome is still incomplete, we first constructing F2 segregation population by individual selfing of D. officinale and D. aduncum F1 population, developing genome-wide SNP markers of the F2 population based on restriction-site associated DNA (RAD) sequencing, and genotying with the whole genome transcriptome sequencing information of Dendrobium species as reference, in order to construct the high density genetic linkage maps; On the other hand, the integrated Dendrobium genetic linkage map will also be constructed based upon our reported primary genetic linkage map results and the new high density genetic linkage maps of D. officinale and D. aduncum. In turn, the comparative genomic study of Dendrobium is expected to be achieved based on the high-density integrated Dendrobium genetic linkage map. The implementation and completion of this project will have contribution to discover the similarities and differences of Dendrobium species from the level of linkage groups (chromosomes). These results also can be used for genome assembing, QTL mapping of significance trait (especial medicinal component), targeted gene cloning and marker-assisted breeding of Dendrobium.

我国是人口大国，也是药用植物储备和需求大国。石斛是名贵的中药材之一，市场需求量大却自然资源紧缺和人工栽培面临种种困难的现状要求我们采取措施积极应对。基于石斛基础研究的现状，以及申请人的工作基础，本项目以构建石斛高密度遗传连锁图谱为起点，进而以通过整合图谱构建开展石斛比较基因组学研究为对象。在考虑石斛基因组仍不完善的前提下，一方面在现有多个石斛F1群体的基础上发展自交F2分离群体，采用简化基因组测序技术，进行群体全基因组SNP发掘和基因型分析，从而构建高密度遗传连锁图谱；另一方面，立足现有的初级遗传连锁图谱，首先进行部分石斛初级整合图谱的绘制，然后综合各类石斛遗传图谱绘制高密度整合图谱，并在此基础上开展石斛比较基因组学研究。本项目的实施和完成将有助于明确石斛不同种间遗传连锁群的异同、指导石斛基因组组装、重要性状（尤其是药用成分）QTL定位、目的基因克隆和标记辅助育种等工作。

高密度遗传图谱是高质量染色体水平基因组组装的重要基础，是开展染色体水平进化和比较基因组学研究的前提。本项目在广泛收集种质资源的基础上，通过构建铁皮石斛和细茎石斛杂交F1代群体，采用SLAF-seq技术对111个作图群体单株进行大规模基因分型，构建了国内外第一张石斛高密度遗传连锁图谱（共8,573个SNP标记，覆盖19个连锁群，总图距为2,737.49 cM，标记间平均图距为0.32 cM）；以石斛茎总多糖含量为首要研究目标，基于该图谱获得相关的QTL位点5个，为开展石斛多糖含量相关功能基因的挖掘找到了候选区段；该图谱还被利用在对现有铁皮石斛全基因组测序结果的组装中，目前完成了822个Scaffold在19条染色体上的挂载（共计约640M的序列，占铁皮石斛全基因组的58%）。本项目不仅开辟了铁皮石斛乃至石斛属基因组学研究的新方向，而且相关研究成果对开展石斛属功能基因学和比较基因组学研究都具有一定的影响。