小麦背景下冰草P基因组抗条锈病基因发掘及初步验证

Stripe rust is one of the most destructive diseases in wheat (Triticum aestivum L.) production. Exploring and identification of novel disease-resistant genes from wild relatives of wheat is an economical and effective strategy to control the prevalence of stripe rust. In our previous study, the intergeneric hybridization of wheat and Agropyron cristatum (A. cristatum) was successfully completed, and then, many additional lines, translocation lines and deletion lines were produced. Further research showed that the locus conferring stripe rust resistance was located on the bin 6PS (0.81-1.00) of translocation line WAT655. Meanwhile, this translocation line was genetically stable, had good comprehensive agronomic characters and was also resistant to leaf rust. In order to clarify the resistance genes underlying stripe rust in this germplasm, resistance gene enrichment and single-molecule real-time sequencing (SMRT RenSeq) and comparative genomics will be applied to identify potential gene sets in the 6PS 0.81-1.00. Candidate genes were screened based on the susceptible mutant, expression patterns and genetic network of these genes under pathogen-induced. Finally, the function of candidate genes was verified by real-time fluorescence quantitative PCR and virus-induced gene silencing. The implementation of this project not only laid the foundation for the cloning of exogenous stripe rust resistance genes from P genome, but also provided technical support for the discovery of genes resistant to leaf rust in this translocation line.

条锈病是危害小麦生产的主要病害之一，而从野生近缘物种中发掘利用抗病新基因是抗病育种的有效途径。前期，我们已经实现了小麦与冰草属间的远缘杂交，并创建了许多附加系、易位系和缺失系。进一步研究表明，条锈病抗性位点被定位于创新种质小麦-冰草易位系WAT655的易位片段6PS 0.81-1.0。同时，此易位系遗传稳定，农艺性状良好，且抗叶锈病。为了阐明该种质中相关的抗条锈病基因，本项目拟首先基于抗性基因富集单分子实时测序（SMRT RenSeq）和比较基因组学，阐明6PS 0.81-1.00区段内潜在的基因集，然后利用EMS诱变筛选感病突变体，确定突变位点或基因，并利用RNA-Seq，确定基因在病原菌诱导下的表达模式和在抗病调控网络中的作用，最终确定抗条锈病候选功能基因，并利用实时荧光定量PCR和病毒诱导基因沉默技术对候选基因的功能进行初步验证。此项目的实施，不仅为冰草外源抗条锈病基因的克隆奠定了基础，还为该区段内抗叶锈病等基因的发掘提供技术支撑。

条锈病是危害小麦生产的主要病害之一，而发掘利用抗病新基因是防治小麦条锈病最经济 有效的方法。冰草（P基因组）是小麦的野生近缘植物，对多种病害具有持久的广谱抗性。前期研究表明，条锈病抗性位点被定位于创新种质小麦-冰草易位系WAT655的易位片段（6PS 0. 81-1.0）。由于，在自然条件下，6PS不能与小麦6号同源群染色体进行配对、重组，且所有已知的冰草品系对条锈病均免疫，因而对其抗病基因进行图位克隆的研究相对困难。本项目基于冰草全长转录组和转录组测序，阐明了6PS 0.81-1.00区段含有32个表达的冰草特异基因，然后根据这些基因在病原菌诱导下的表达模式，通过特异分子标记开发和冰草BAC文库筛选、测序和组装，共克隆了4个冰草抗条锈病候选基因和6个基因的CDS。转基因功能验证表明了Agr6971、Agr4080和Agr8173的转基因植株能够达到抗和中抗小麦条锈病的水平，是创新种质小麦-冰草易位系WAT655的易位片段的抗病遗传基础。此项目的实施，不仅为冰草外源抗条锈病基因的克隆奠定了基础，还为小麦背景下外源抗病基因的发掘提供了新思路。