华北落叶松结实性状相关EST-SSR标记的挖掘

Larix principis-rupprechtii is one of native tree species in North China. However, low seed production in seed orchard and long breeding cycle became the primary concern during its improvement program, the same as other species in Larix genus. Therefore, it is urgent to explore some new strategies or methods to make clear the reproduction genetic nature, which would improve the breeding program. Molecular marker-assisted selection is the most powerful and effective approach that can accelerate breeding progress because it can identify genotypes efficiently during plant juvenile stage. However, there are few data on molecular marker-assisted selection of seed production in Larix genus. On the basis of seed production investigation, progeny test and EST-SSR exploration in Larix principis-rupprechtii, a large number of EST-SSR markers will be firstly developed from De novo assembly transcriptome sequences of Larix principis-rupprechtii tissues or organs including strobilus, branch, needle, inflorescence etc. Then these markers will be screened against the predominantly male and female pools. Finally, the screened candidate markers above will be used to associate with 8 important reproduction traits in 138 Larix principis-rupprechtii half-sib based on their population structure and individual kinship relationship. This project will have a strong impact on molecular marker-assisted selection in Larix breeding program in the future.

华北落叶松是我国特有的落叶松属树种之一，种子园产种量低和育种周期长是制约其遗传改良成果应用的关键因素，急需探索新技术或新方法来揭示结实量的遗传本质来提高落叶松的育种水平。分子标记辅助选择育种技术因能实现对目标性状的早期选择，缩短育种年限而被认为是最强有力的方法。鉴于落叶松结实性状无相关报道，课题组将在前期结实及子代测定和探索EST-SSR开发的基础上率先深入开展此方面研究。首先对华北落叶松多个组织或器官进行高通量RNA-seq测序；而后利用生物信息学方法批量开发EST-SSR标记，并通过BSA法混合偏雄和偏雌两个DNA集群池进行候选标记筛选；最后以138份华北落叶松半同胞子代为关联群体，在分析群体结构和亲缘关系的基础上，利用关联分析对候选EST-SSR标记和8个与偏雄/偏雌相关的结实性状进行联合分析寻找相关位点并挖掘优异等位基因。本研究的实施将为实现落叶松分子标记辅助选择育种奠定基础。

华北落叶松是我国特有的落叶松属树种之一，种子园产种量低和育种周期长是制约其遗传改良成果应用的关键因素，急需探索新技术或新方法来揭示结实量的遗传本质来提高落叶松的育种水平。分子标记辅助选择育种技术因能实现对目标性状的早期选择，缩短育种年限而被认为是最强有力的方法。鉴于落叶松结实性状无相关报道，课题组将率先开展此方面研究。首先对华北落叶松雌雄花芽进行高通量RNA-seq测序；而后利用生物信息学方法批量开发了588对EST-SSR标记，并通过BSA法混合偏雄和偏雌两个DNA集群池进行候选标记的筛选，挑选出了多态性标记106对；最后以138份华北落叶松半同胞子代为关联群体，利用关联分析对效果最佳的40对候选EST-SSR标记和与偏雄/偏雌相关的结实性状进行联合分析寻找相关位点并挖掘优异等位基因。主要研究结果如下：（1）建立了完整的华北落叶松结实表型数据库，性状之间存在丰富的遗传变异，构建了包括138个无性系在内的结实性状关联群体。（2）测定了138个半同胞家系关联群体的子代生长性状。（3）率先开展华北落叶松转录组测序，拼接后共得到124204条Unigenes，功能注释显示共有约56761（45.7%）条可分别在SWISS-PROT、TREMBL、PFAM、NR和 KOG等数据库中得到注释；另外还首次找到与落叶松偏雌相关的147条Unigenes。（4）基于华北落叶松转录组拼接序列批量鉴定以及合成588对EST-SSR引物，并通过偏雌和偏雄BSA混合池鉴定出106对潜在的候选多态性EST-SSR引物。（5）从分子标记水平证实当前138份华北落叶松关联群体具有中等水平遗传多样性，针对落叶松自交不亲和现象，提出了可提高育种效率的亲本选择策略。本研究的实施将为我国实现落叶松分子标记辅助选择育种奠定基础。