Characterisation of major genes (Stb)-mediated resistance to Septoria tritici blotch disease in wheat

小麦是最重要的主食作物之一，提供全球人类每日总热量消耗的20%。三孢壳针孢斑点病（STB）是在世界上大多数小麦种植区遇到的毁灭性疾病，并且是英国小麦的主要叶部疾病，尽管进行了杀真菌剂处理，但仍造成每年5-10%的小麦损失。STB的病原体是真菌Zymoseptoria triumph。真菌群体中杀菌剂抗性的产生和蔓延严重威胁着小麦生产和粮食安全，因此抗STB是小麦育种的重要目标。到目前为止，21个主效（Stb）基因和几个数量性状位点（QTL），使较小的贡献，抗性表型已被确定和遗传作图。到目前为止，这些基因中只有一个被克隆，抗性机制仍然知之甚少。由于缺乏具有加性效应的明确的QTL和几乎没有诊断标记，目前的STB抗性育种策略主要依赖于育种材料的表型评价，而不是基于靶向基因分型的选择。因此，在过去的几十年里，只有一个温和的改善STB抗性的商业小麦种质和大多数目前AHDB推荐的英国小麦品种只显示中等抗性STB。发表的遗传图谱在Stb位点大多是低分辨率，往往涉及早代标记（如RFLP，SSR），这被认为是不切实际的现代分子标记辅助育种。目前尚不清楚是否有任何Stb基因（Stb 6除外）可能存在于英国商业小麦中，或者这些基因在英国条件下控制STB的有效性如何。该博士项目旨在填补小麦育种研究中的这一重大空白，通过解决以下目标来提高我们对已知Stb基因控制的STB抗性的遗传学和机制的理解：1.确定目前英国田间Z.评估抗性是否在幼苗和/或成株阶段起作用3.精细定位最感兴趣和有用的抗性基因，并开发用于育种的诊断性第二代遗传标记（KASP）4.深入了解Stb基因如何Stb基因将通过杂交从外来和外来基因型转移到高度易感的英国小麦背景中。绘制种群近等基因系的工作已经在进行中。然后将使用当前UK字段Z对这些材料进行表型分析。在温室条件下，随后进行高密度基因分型。我们最近克隆了Stb 6，并表明它编码一种Wall相关受体样激酶（WAK）蛋白，该蛋白定义了一类新的抗病蛋白（Nat Genet，已接受）。使用最新的小麦基因组IWGSC组装，我们已经注释并手动策划了所有WAK编码基因，并假设其中至少有一些可能对应于Stb基因。事实上，生物信息学分析表明，几个Stb基因座对应于含有一个或多个WAK基因的物理间隔。学生将通过使用我们实验室建立的病毒诱导基因沉默（VIGS）方法（植物生理学160：582-90）评估候选WAK基因的功能来进一步测试上述假设。将使用分子生物学和生物成像方法的组合来研究Stb基因赋予的抗性机制，特别是确定抗病性是否与细胞死亡相关，真菌生命周期中的哪一步被靶向，以及激活了哪些防御途径。所获得的知识将有助于抗STB育种，从而有助于提高作物产量。

Wheat is one of the most important staple food crops providing 20% of total daily calories consumed by humans worldwide. Septoria tritici blotch (STB) is a devastating disease encountered in most wheat growing areas of the world and is the primary foliar disease of wheat in the UK responsible for year-on-year wheat losses of 5-10% despite fungicide treatments. The causal agent of STB is the fungus Zymoseptoria tritici. Emergence and spread of fungicide resistance in fungal populations seriously threatens wheat production and compromises food security, therefore resistance to STB is an important target in wheat breeding. To date, 21 major (Stb) genes and several quantitative trait loci (QTL) that make smaller contributions to the resistance phenotype have been identified and mapped genetically. Only one of these genes has so far been cloned and mechanisms of resistance remain poorly understood. Owing to a lack of well-defined QTLs with additive effects and near absence of diagnostic markers, the current STB resistance breeding strategies rely mostly on phenotypic evaluation of breeding materials rather than targeted genotyping based selection. Consequently, over the past decades, there has been only a modest improvement of STB resistance in the commercial wheat germplasm and the majority of current AHDB recommended UK wheat cultivars show only moderate resistance to STB.Published genetic maps at Stb loci are mostly low resolution and often involve early generation markers (eg RFLP, SSR), which are considered impractical in modern marker-assisted breeding. It is not known whether any of the Stb genes (apart from Stb6) may be present in the UK commercial wheat, or how effective these may be in controlling STB under UK conditions. The proposed PhD project, aims to fill this significant void in wheat breeding research by enhancing our understanding of genetics and mechanisms of STB resistance controlled by known Stb genes through addressing the following objectives:1.Identify frequency of virulence/avirulence in the current UK field Z. tritici populations towards each Stb gene, and whether any of these genes confer broad-spectrum resistance2.Assess whether resistance operates at seedling and/or adult plant stage3.Fine-map the most interesting and useful resistance gene(s), and develop the diagnostic second generation genetic markers (KASP) for use in breeding4.Gain insight into how the Stb gene(s) are conferring resistanceThe Stb genes will be transferred by crossing from foreign and exotic genotypes into a highly susceptible UK wheat background. Development of mapping populations nearly-isogenic lines is already in progress. These materials will then be phenotyped with current UK field Z. tritici isolates under glasshouse conditions followed by high-density genotyping. We have recently cloned Stb6 and shown that it encodes a Wall-associated receptor-like kinase (WAK) protein that defines a novel class of disease resistance proteins (Nat Genet, accepted). Using the latest IWGSC assembly of wheat genome, we have annotated and manually curated all WAK encoding genes and hypothesised that at least some of these may correspond to Stb genes. Indeed, bioinformatics analyses have indicated that several Stb loci correspond to physical intervals containing one or more WAK genes. The student will further test the above hypothesis by assessing the function of candidate WAK genes using Virus-induced gene silencing (VIGS) approach well established in our laboratory (Plant Physiol 160: 582-90). Mechanisms resistance conferred by the Stb genes will be investigated using a combination of molecular biology and bioimaging approaches, in particular determining whether disease resistance is associated or not with cell death, what step in the fungal lifecycle is being targeted, and what defence pathways are activated. The knowledge gained will aid breeding for STB resistance and therefore help to enhance crop productivity.