高粱抗丝黑穗病基因精细定位及候选基因鉴定

Fine mapping is the basis of gene discovery for important agronomic traits and for molecular marker identification. Sorghum head smut is one of the main diseases in sorghum production. However, sorghum resistant gene to head smut is not precisely mapped enough at current time, so its gene cloning and molecular marker identification are highly restricted. During our early study, we conducted genome-wide association study on RAD-seq for 415 sorghum varieties , and we mapped the SNP locus as related to sorghum resistant gene to head smut. For finer mapping results, based on RAD-Seq technology, recombinant inbred line ( RIL) was proposed to adopt in material structuring, as resistant to sorghum head smut for this project, so that head smut resistant gene of Sorghum bicolor was mapped furthermore. Then we compared those results with GWAS mapping results to define initial mapping section, as precise as possible for resistant gene to head smut. Then we selected disease-resistant individuals with backgrounds similar to those of susceptible parent plants, for we prepare to backcross and establish the colony for fine mapping, and then we developed amplified fragment length polymorphism molecular marker, screened the recon fine mapping for sorghum resistant genes to head smut. Next, we made use of sequence variation of the candidate genes from anti-infectious materials in the fine mapping section. Meanwhile we employed anti-infectious materials with different genetic backgrounds to estimate the practicability of fine mapping for molecular markers. Fine mapping results for sorghum resistant gene to head smut were to be obtained in this study, and this would lay a foundation for molecular breeding and molecular mechanism study on sorghum resistance to head smut.

精细定位是重要农艺性状基因挖掘和分子标记鉴定的基础。高粱丝黑穗病是影响高粱生产安全的主要病害，但目前高粱抗丝黑穗病基因定位不够精准，限制了其基因克隆和分子标记的鉴定。在前期研究中，我们利用415份高粱品种简化基因组测序的全基因组关联分析，定位了与高粱抗丝黑穗病相关的SNP位点。为获得更精细定位结果，本项目拟利用高粱抗感丝黑穗病材料构建的重组自交系基于简化基因组测序技术进一步定位高粱抗丝黑穗病基因，与GWAS定位结果比较，获得精准的抗丝黑穗病初定位区间，再从RIL中选择与感病亲本背景相似的抗病个体回交构建精细定位群体，开发多态性分子标记，筛选重组子精细定位高粱抗丝黑穗病基因。然后通过精细定位区间内的候选基因在抗感材料中序列变异筛选候选基因，并利用不同遗传背景抗感材料评价精细定位分子标记的实用性。本研究将获得高粱抗丝黑穗病基因的精细定位结果，也将为高粱抗丝黑穗病分子育种和分子机制研究奠定基础。

精细定位是重要农艺性状基因挖掘和分子标记鉴定的基础。高粱丝黑穗病是影响高粱生产安全的主要病害，但目前高粱抗丝黑穗病基因定位不够精准，限制了其基因克隆和分子标记的鉴定。我们利用428份高粱品种简化基因组测序的全基因组关联分析，定位了与高粱抗丝黑穗病相关的SNP位点。428份材料经多年长期接种高粱丝黑穗病菌3号生理小种鉴定获得了准确表型数据，免疫品种14份、高抗品种33份、抗品种20份、中抗品种28份、感病品种163份、高感品种170份。为获得更精细定位结果，本项目利用高粱抗感丝黑穗病材料构建的重组自交系基于简化基因组测序技术进一步定位高粱抗丝黑穗病基因，根据关联阈值判定，共得到2个区域，总长度为35.49 Mb，共包含591个基因，其中非同义突变SNP位点的基因共186个。与GWAS定位结果比较，获得精准的抗丝黑穗病初定位区间，两种方法高粱抗丝黑穗病基因均定位在同一条染色体上，中间遇到着丝点，前端17.7Mb，后端29.5Mb两个区间，总区段大小在55.7Mb范围内。再从RIL中选择与感病亲本背景相似的抗病个体回交构建精细定位群体，获得800个单株基因型。开发初定位区间内抗感亲本多态性标记283个，筛选了纯合抗重组体15个，根据重组体抗丝黑穗病鉴定结果精细定位抗丝黑穗病基因区间缩短为前端1.8Mb和后端2.3Mb范围内。根据精细定位范围内，两段区间有37个候选基因。根据基因注释有3个相关候选基因有可能参与高粱抗丝黑穗病抗性机制，其中1个在亲本间有序列差异，但在抗感品种后代中不是100%正确，因此，未筛选到高粱抗丝黑穗病候选基因。精细定位区间开发的多态性15个SSR标记，在后代抗感群体中比较分析有1个标记6Jsmt367可在育种中尝试，但由于鉴定抗感品种量不够大，仍需要严谨进一步去分析验证其在育种中的实用价值。本研究获得了高粱抗丝黑穗病基因的精细定位结果，这将为高粱抗丝黑穗病分子育种和抗病机制研究奠定理论基础。