玉米第1染色体抗灰斑病主效QTL精细定位与候选基因发掘

At present, maize gray leaf spot (GLS), caused by the fungus Cercospora zeae-maydis, is recognized as one of the most significant yield-limiting diseases. The most economical and effective way to control this disease is the utilization of resistant varieties, but maize breeding for gray leaf spot (GLS) resistance has been hindered by the quantitative nature of the inheritance of GLS resistance and by the limitations of selection under less than optimum disease pressure. So it possesses significant importance to digging GLS resistant genes. Although a number of research efforts had been conducted on GLS resistant gemplasm selection and QTL mapping, the number and effect of genes involved in GLS resistance are still not clear. As a quantitative trait, it's necessary to conduct fine mapping on major QTL for understanding its genetic basis and further gene clones. In our previous study, a pair of introgression lines that display difference in major QTL region on chromosome1 of maize were developed at Shenyang, being derived from a cross of 81162(susceptible) as the recurrent parent and CN165 (resistant) as the donor parent, followed by two backcross cycles and finished with two to three generations of inbreeding by artificially inoculating Cercospora zeae-maydis in the field. In this study, the introgression lines were used as materials. The GLS major QTL region is firstly fine-mapped by polymorphic markers which come from published maize genome sequencing information with a secondary segregation population derived from the introgression lines. Furthermore, according to the precise positioning results, using bioinformatics methods to explore positional candidate genes, and its functional SNP and haplotype will be identified using a linkage and association mapping with GLS resistant introgression lines and inbred lines. The results of this study provide useful information on understanding GLS genetic mechanism and its usage on molecular assisted selection.

当前，尾孢菌引起的玉米灰斑病严重影响着玉米生产，防治该病害最经济有效的途径是采用抗病品种，因此抗灰斑病基因发掘具有重要意义。玉米抗灰斑病为数量性状，要精确地了解该性状的遗传基础并实现QTL的克隆，须对主效QTL进行精细定位。本课题组以抗病自交系CN165为供体，感病自交系81162为受体，经多代回交自交并结合人工接种鉴定获得了玉米第1染色体主效QTL区段存在差异的一对导入系。本研究将以这一对导入系为试材，通过杂交自交构建次级分离大群体，并利用已公布的玉米全基因组测序信息，筛选QTL区段内所有的多态性标记，对分离群体单株基因型检测，寻找目标染色体区域内发生交换的单株，进行QTL精细定位；根据精细定位结果，利用生物信息学手段发掘位置候选基因；借助自然群体和回交导入系的标记和表型关联分析，确定候选基因区段内的功能性差异SNP和单体型。研究结果有助于解析玉米灰斑病的抗性遗传基础和进行标记辅助育种。

由尾孢菌侵染玉米叶片引起的玉米灰斑病已成为我国东华北及西南玉米产区重要的病害之一。其中引起东北地区灰斑病的病原菌为玉蜀黍尾孢菌，引起西南地区灰斑病的病原菌为玉米尾孢菌（刘文奎等，2013）。该病在整个生育期皆可发生，但以抽雄和灌浆期发病为重。发病植株由于叶片损伤和凋萎使光合组织丧失功能，最终引起穗粒数减少，粒重降低，产量损失在10%-70%之间。培育和种植抗病品种是防治灰斑病最经济有效的方法。.抗源材料，是抗病品种选育工作的关键。研究表明，拥有亚热带或热带血缘的 PB 类玉米种质，尤其是78599后代选系，具有相对较好的抗性。本项目分别以抗病自交系CN165和齐319为供体，感病自交系81162和掖478为受体，经多代回交自交获得的回交导入系为试材，对第1染色体上抗灰斑病QTL进行精细定位。并借助基因组序列信息，发掘玉米抗灰斑病候选基因。研究结果表明，不同供体材料中第1染色体上的主效抗病QTL位置不同，其中CN165材料中主效抗病QTL位于染色体1.05-1.06区间，齐319中主效抗病QTL位于1.02-1.03区间。将导入系与轮回亲本回交和自交创造目标染色体区间的重组体，通过对重组体基因型和抗病表现型分析，将CN165中抗病QTL定位于标记snym22和SYLY-29之间，标记间距离为4.35Mb；将齐319中抗病QTL定位于标记FFF4G和Fmap6B之间，标记间物理距离为363.61Kb。借助201个自交系的自然群体标记和表型关联分析，标记SYLY-80可以有效用于抗病基因选择。进一步对FFF4G和Fmap6B区间内序列信息进行分析，确定一个候选基因，经克隆该基因在齐319和掖478以及B73之间存在较大差异，且在抗感材料间表达差异显著。该研究说明玉米第1染色体上存在较多的抗病基因，研究结果为第1染色体上抗病基因的克隆奠定了基础，对培育抗病品种具有重要意义。