【作者】 朱晓彪；

【导师】 王秀娥；

【作者基本信息】 南京农业大学 ， 作物遗传育种， 2011， 博士

【摘要】 小麦黄花叶病(Wheat yellow mosaic, WYM)是由小麦黄花叶病毒(Wheat yellow mosaic bymovirus, WYMV)引起的一种土传病毒病害,在冬小麦种植区经常发生,正日益成为危害我国小麦生产的最严重的病害之一,危害小麦安全生产。抗WYMV基因资源的挖掘和深入研究对于小麦抗病毒育种具有重要意义。‘西风小麦’是自日本引进的早熟抗病优良品种,具有抗倒伏、白粉病、赤霉病、条锈病、穗发芽和高抗WYMV等优异性状。在我国,育种工作者利用‘西风小麦’作为骨干亲本直接或间接育成了一些广泛推广的优良品种,如‘宁麦9号’、‘宁麦16号’和‘扬麦18号’,这些品种也高抗WYMV.‘镇9523’是江苏丘陵地区镇江农科所育成的综合性状优良的品种,具有矮秆茎粗、穗大粒多、粒重高、粒型好,中抗赤霉病、叶锈病、纹枯病等优异性状,但高感WYMV.本研究以‘西风小麦×镇9523’重组自交系群体构建了分子标记连锁图谱,在此基础上,结合WYMV抗性、株高、穗长和每穗小穗数等4个性状的考察和鉴定,进行QTL分析,为小麦育种提供新的基因资源。研究还针对鉴定出的一个抗WYMV主效QTL QYm.nau-5A.1,构建次级F2分离群体进行精细定位,为图位克隆分离该主效QTL奠定重要的基础。1.抗WYMV QTL分析和主效QTL QYm.nau-5A.1的精细定位应用植物数量性状主基因+多基因混合遗传模型对WYMV抗性进行遗传分析,结果表明,WYMV抗性的遗传在四个试验环境中符合2或3对主基因+多基因混合遗传模型,主基因的遗传率为81.00-93.93%,多基因的遗传率为5.55-17.17%。利用覆盖小麦全基因组的1790对SSR、STS、EST-SSR和EST-STS标记在‘西风小麦×镇9523’RIL群体双亲之间进行多态性筛选,选取在双亲及群体中多态性较好且带型清晰的317个标记,利用JoinMap 4.0软件进行分子标记连锁图谱的构建,271个标记的274个位点组成了33个连锁群,连锁群总长度为1685.3cM,标记位点间的平均距离为9.8cM,覆盖小麦的21条染色体。在此基础上,结合群体四个环境的WYMV抗性鉴定结果,利用Windows QTL Cartographer V2.5软件的复合区间作图法对WYMV抗性进行QTL定位,共检测到3个新的抗WYMV QTL:QYm.nau-3B.1、QYm.nau-5A.1和QYm.nau-7B.1,分别位于染色体3BS、5AL和7BS上。抗病等位基因均来自抗病亲本‘西风小麦’。其中QYm.nau-3B.1和QYm.nau-5A.1在四个环境中均被检测到,分别解释3.3-10.2%和25.9-53.7%的表型变异；而QYm.nau-7B.1仅在试验环境E1中被检测到,解释4.9%的表型变异。根据抗WYMV QTL分析的结果,在RIL群体中选择一个仅含有QYm.nau-5A.1且其他农艺性状和镇9523相似的高抗家系与镇9523杂交,构建了一个包含6002个单株的次级F2分离群体来精细定位QYm.nau-5A.1,经重组体筛选将QYm.nau-5A.1定位于标记Xwmc415.1和5EST-440之间,且2个EST-STS标记5EST-44和5EST-90与QYm.nau-5A.1表现出共分离。连锁分析表明,标记Xwmc415.1、5EST-44和5EST-90与QYm.nau-5A.1之间的遗传距离均为0.0cM,标记5EST-440与QYm.nau-5A.1之间的遗传距离为O.1cM。最终将QYm.nau-5A.1定位于0.1cM的标记区间Xwmc415.1-5EST-440内,为图位克隆分离QYm.nau-5A.1奠定了的基础。利用与QYm.nau-5A.1紧密连锁的3个标记Xwmc415.1、5EST-44和5EST-90对WYMV抗性已知的46个品种组成的小麦品种群体进行分子标记分析,结果在10个抗WYMV品种中检测到QYm.nau-5A.1,且这3个标记在所有12个感WYMV品种中均扩增出和感病品种‘镇9523’一致的与感病相关的带型。因此,这3个标记在小麦品种群体中能够对QYm.nau-5A.1进行有效地鉴定,可用于抗WYMV MAS育种。2.三个重要农艺性状的QTL分析应用植物数量性状主基因+多基因混合遗传模型对株高、穗长和每穗小穗数等3个农艺性状进行遗传分析,结果表明,株高、穗长和每穗小穗数性状的遗传在两个试验环境中均符合2对主基因+多基因混合遗传模型。在构建分子标记连锁图谱的基础上,结合群体两个环境目标性状的考察结果,利用Windows QTL Cartographer V2.5软件的复合区间作图法对3个农艺性状进行QTL定位,共检测到4个与株高相关的QTL:QPh.nau-2D、QPh.nau-3B.1、QPh.nau-4B和QPh.nau-4D,分别位于染色体2DS、3BL、4BL和4DS上。其中QPh.nau-2D, QPh.nau-4B和QPh.nau-4D在两个环境中均被检测到,可分别解释7.4-7.9%、29.3-30.3%和28.3-35.6%的表型变异。共检测到5个与穗长相关的QTL:QSl.nau-2D.α、QSl.nau-2D.b, QSl.nau-5A.1、QSl.nau-5B和QSl.nau-6B,分别位于染色体2DS、2DS、5AL、5BS和6BL上。其中,QSl.nau-2D.α、QSl.nau-2D.b和QSl.nau-5A.1在两个环境中均被检测到,可分别解释20.6-29.0%、5.0%和8.8-11.6%的表型变异。共检测到5个与每穗小穗数相关的QTL:QSn.nau-1A.1、QSn.nau-5A.1.α、QSn.nau-5A.1.b、QSn.nau-5D和QSn.nau-6B,分别位于染色体1AS、5AL、5AL、5DL和6BL上。其中,QSn.nau-1A.1和QSn.nau-5D在两个环境中均被检测到,分别解释10.2-11.1%和7.8%-11.9%的表型变异。更多还原

【Abstract】 Wheat yellow mosaic (WYM), which caused by wheat yellow mosaic bymovirus (WYMV) is a soil-borne bymovirus disease, is an important disease in the winter wheat growing regions and it has growing as one of the most serious diseases in wheat production of China. The mining of more resistance gene resources is of critical importance in wheat breeding for WYMV resistance.The wheat variety,’Xifeng Wheat’, originally introduced from Japan into China, possesses many elite characters such as resistances to powdery mildew, fusarium head blight, stripe rust, WYMV as well as resistances to lodging and pre-harvest sprouting. Several new wheat varieties with high level of resistance to WYMV have been released using’Xifeng Wheat’as breeding parents directly or indirectly in China (e.g.’Ningmai 9’, ’Ningmai 16’and’Yangmai 18’). The wheat variety,’Zhen 9523’, developed by Jiangsu Hilly Area Zhenjiang Agricultural Research Institute, strong stem, bigger spikes and more grains, higher grain weight, good grain quality, and has moderately resistant to fusarium head blight, leaf rust and sheath blight, but it is highly susceptible to WYMV. In the present research, a RIL population was constructed using the two varieties as parents, and a molecular marker based linkage map was established. Based on the evaluation of WYMV resistance, QTLs associated with WYMV resistance were analyzed. A secondary F2 population was constructed to fine map a major QTL QYm.nau-5A.1 identified for WYMV resistance in the RIL population, which would provide an important foundation for the map-based cloning of QYm.nau-5A.1. The major results obtained were as following:1. Identification of QTLs for WYMV resistance in the RIL populationGenetic analysis for the WYMV resistance was conducted by mixed major gene plus polygene inheritance models. Results showed that the WYMV resistance was controlled by two or three major genes plus polygenes, major genes and polygenes heritabilities were 81.00-93.93% and 5.55-17.17%, respectively. Among the 1,790 SSR, STS, EST-SSR and EST-STS markers,317 primer pairs could amplify clear and reproducible polymorphic band (s) between the two parents and the RILs. Among them, when using the JoinMap 4.0 software,271 markers representing 274 non-redundant loci assigned to the molecular marker-based linkage map contained 33 linkage groups spanning a total genetic length of 1,685.3cM with an average marker interval of 9.8cM. All the 21 wheat chromosomes were represented by at least one linkage group. Based on the molecular marker-based linkage map and phenotypic data of WYMV resistance collected from four-year, two-location replicated field trials, QTLs associated with the WYMV resistance were detected with the Windows QTL Cartographer V2.5 software by composite interval mapping (CIM) method. Three QTLs, QYm.nau-3B.1, QYm.nau-5A.1 and QYm.nau-7B.1, were detected on chromosomes 3BS,5AL and 7BS, respectively. The increased allele effects were all provided by the resistant parent’Xifeng Wheat’. Among the three QTLs, QYm.nau-3B.1 and QYm.nau-5A.1 were detected in all the four trials, and could explain 3.3-10.2% and 25.9-53.7% of the phenotypic variation, respectively, while QYm.nau-7B.1 was detected in one trial that explained 4.9% of the phenotypic variation.2. Fine mapping of the major QTL QYm.nau-5A.1In order to fine map the QYm.nau-5A.1, based on QTL analysis for WYMV resistance, a secondary F2 population consisting 6002 plants were constructed by a cross ’RILV-6’and ’Zhen 9523’, and the highly WYMV-resistant RIL’RILV-6’only contained QYm.nau-5A.1 has similar agronomic traits with’Zhen 9523’. When using the markers to screen the recombinats in all the 1340 susceptible plants, the QYm.nau-5A.1was flanked by Xwmc415.1 and 5EST-440, and the two EST-STS markers were co-separated with the QYm.nau-5A.1. Linkage analysis showed that the genetic distances were 0.0cM between Xwmc415.1,5EST-44,5EST-90 and QYm.nau-5A.1, and that the genetic distances were 0.1 cM between 5EST-440 and QYm.nau-5A.1. Finally the QYm.nau-5A.1 was flanked by the two markers Xwmc415.1 and 5EST-440 with a distance of 0.1cM, which would provide an important foundation for the map-based cloning of QYm.nau-5A.1.Three markers, Xwmc415.1,5EST-44 and 5EST-90 closely linked with QYm.nau-5A.1 identified in the present study, were used for marker analysis of the wheat varieties population consisting of 46 varieties with known WYMV resistance. Result showed that that QYm.nau-5A.1 was present in 12 varieties with WYMV-resistance, and all the 12 WYMV-susceptible varieties amplified the same specific band as in’Zhen 9523’using the three markers. So the three markers were very effective to identify the QYm.nau-5A.1 in the wheat varieties population, and they should be useful in marker-assisted selection (MAS) of WYMV resistance in wheat breeding.3. QTL analysis of the three important agronomic traits in the RIL populationGenetic analysis for plant height, spike length and spikelet number per spike were conducted by mixed major gene plus polygene inheritance models. Results showed that the three agronomic traits were all controlled by two major genes plus polygenes.Based on the molecular marker-based linkage map and phenotypic data of the three agronomic traits collected from two-year, one-location replicated field trials, QTLs associated with the three traits were detected with the Windows QTL Cartographer V2.5 software by CIM method. Four QTLs for the plant height, QPh.nau-2D, QPh.nau-3B.1, QPh.nau-4B and QPh.nau-4D, were detected on chromosomes 2DS,3BL,4BL and 4DS, respectively. Among them, QPh.nau-2D, QPh.nau-4B and QPh.nau-4D were all detected in all the two trials and could explain 7.4-7.9%,28.3-35.6% and 29.3-30.3% of the phenotypic variation, respectively. Five QTLs for the spike length, QSl.nau-2D.a, QSl.nau-2D.b, QSl.nau-5A.1, QSl.nau-5B and QSl.nau-6B, were detected on chromosomes 2DS,2DS, 5AL,5BS and 6BL, respectively. Among them, QSl.nau-2D.a, QSl.nau-2D.b and QSl.nau-5A.1 were detected in all the two trials and could explain 20.6-29.0%,5.0% and 8.8-11.6% of the phenotypic variation, respectively. Five QTLs for the spikelet number per spike, QSn.nau-1A.1, QSn.nau-5A.1.a, QSn.nau-5A.1.b, QSn.nau-5D and QSn.nau-6B, were detected on chromosomes 1AS,5AL,5AL,5DL and 6BL, respectively. Among them, QSn.nau-1A.1 and QSn.nau-5D were detected in all the two trials and could explain 10.2-11.1% and 7.8-11.9% of the phenotypic variation, respectively.更多还原