【作者】 赵志杰；

【导师】 田保明； 铁双贵；

【作者基本信息】 郑州大学 ， 作物遗传育种， 2013， 硕士

【摘要】 玉米(Maize)是当今社会中所发挥的作用越来越大,它不仅仅是重要的粮食作物,也是重要的饲料及工业原料。我国的玉米总产量仅次于水稻产量,在国民经济中占有越来越重要的地位。玉米粗缩病是由玉米粗缩病病毒引起的一种世界性病害,也是我国黄淮海玉米产区的重要病害之一。在我国引起玉米粗缩病的病原为水稻黑条矮缩病毒(RBSDV),传毒媒介为灰飞虱。玉米粗缩病发病植株显著矮化、多数不抽雄、雌穗不结实或籽粒少、产量损失严重甚至绝收。为了控制病害流行,减少生产损失,培育出抗性良种并在现有品种基础上进行种质的遗传改良是防治该病的有效方法。本文系统介绍了玉米粗缩病的病原、病毒寄主、病毒传播方式、危害症状、发病规律以及抗性鉴定方法等,构建了室内人工饲养灰飞虱并用于对玉米材料进行传毒接种的技术平台,并在此基础上,进行了以下三方面的研究：(1)对室内人工饲养扩繁无毒灰飞虱的技术条件进行了探索,分别研究了不同温度(21℃、23℃、25℃、27℃和29℃)对灰飞虱成活率的影响和对灰飞虱产卵量的影响,结果表明,灰飞虱在23℃时成活率最高,21-29℃范围内,随温度逐渐升高,灰飞虱卵孵化量呈递增趋势。最终确定了室内扩繁灰飞虱的最适温度为25。C±2℃。(2)对3个不同地区(连云港、原阳和保定)采集的发病玉米、水稻和小麦植株等5个样本进行RBSDV病毒核酸提取,对其S9片段测序后进行序列比对,结果显示5个样本的同源性为97.92%,27处突变碱基中有1处造成氨基酸理化性质改变,且突变位点位于第一个开放阅读框。原阳水稻和玉米中RBSDV病毒样本与连云港水稻中RBSDV病毒样本核酸序列同源性高于河北保定小麦和玉米的RBSDV病毒样本。(3)对郑58(感)、C7-2(中抗)、Mo17(感)、478(高感)、丹340(抗)、P138(高抗)、郑单958(感)和XH6(未知)等八个不同抗性的玉米材料进行不同带毒率灰飞虱的RBSDV人工接种,并利用荧光定量RT-PCR技术研究玉米材料接种RBSDV后,植株体内病毒增殖动态变化。研究结果如下：对灰飞虱分别进行12h、24h、48h、72h和120h饲毒后,灰飞虱的带毒率分别为10%、13.3%、233%、30%和30%。通过比较不同处理的材料平均含毒量发现,灰飞虱带毒率越高,玉米植株体内病毒含量越高,发病越快,症状越严重。其中带毒率23.3%的处理中各材料发病情况较理想,平均病毒最大含量和最小含量的差异也最大,能够更好地鉴定出材料的抗病差异性。5个不同带毒率的灰飞虱处理分别进行玉米接种传毒后,10%、13.3%带毒率的处理中发病不均一,30%带毒率的处理中材料发病过重,部分植株过早死亡。带毒率23.3%的处理中各材料发病均一,能较好地鉴定出材料的抗性水平。P138、C7-2和丹340的抗性水平较其他材料好,其他感病的材料病毒含量数量级均在106copy/μl。在带毒率为10%的传毒处理中,某些材料检测到少量病毒但并不发病。所有玉米材料的初始发病时间均在四叶一心期,即开始接种后的第28天观察到发病症状。抗病材料接毒后体内病毒增殖慢,且病毒峰值极显著小于高感材料。感病材料接毒后,体内病毒近似呈线性增殖,病毒繁殖快,且峰值高。更多还原

【Abstract】 Maize is important in the world that can be used for food, feed and industrial materials. In our country, the maize yield ranking only second to rice, occupies a very important position in the national economy. Maize dwarf rough disease is a worldwide disease, and is one of the important diseases of maize in Huang-Huai-Hai region of China. In China, the disease is caused by Rice Black-streaked Dwarf Virus (RBSDV), which transmission vector is Laodelphax striatellus Fallen. Susceptible plants are dwarf significantly, no tassel, no grains, yield losses, and even have no harvest. In order to control disease epidemics, and reduce yield loss, breeding and genetic improvement is the most economic and effective ways of controlling the disease.This study makes a systematic introduction on pathogen of MRDD, host, spread way, symptoms, occurrence regularity, and resistance identification method, build technology platform of indoor artificial propagate Laodelphax striatellus Fallen, and artificial inoculation of virus. And then conduct the following three researches:1.Explore the technology conditions of indoor artificial propagate Laodelphax striatellus Fallen. Study the influence of survival rate and oviposition amount of Laodelphax striatellus Fallen in different temperature (21℃, and23℃, and25℃, and27℃,and29℃). The results indicate that high survival rate of laodelphax striatellus at23℃, the eggs hatch volume increment within21℃-29℃. Finally,25℃±2℃is choosed as the optimum temperature of indoor propagation of laodelphax striatellus.2. Choose3different area (Lianyungang, and Yuan yang and Baoding) diseased maize, rice and wheat plant,5samples for RBSDV virus nucleic acid extraction, sequence its S9measuring and blast in NCBI. Results show that5samples of homologous is97.92%,2of27bases mutations cause amino acids physic-chemical nature changed and are located in the first ORF. The nucleic acid sequence homology of RBSDV samples in Yuan yang and Lianyungang over samples in Hebei Baoding.3. In order to study the virus development in infected maize plant,8different maize materials(Zheng58, and C7-2, and Mo17, and478, and Dan340, and P138, and Zhengdan958and XH6) were detected after artificial inoculation by laodelphax striatelluswith different carrier rate. Three virus-acquisition time of laodelphax striatellus were settled as12h,24h,48h,72h,120h respectively, and the carrier rate were10%,13.3%,23.3%,30%,30%.Using Real-time fluorescence quantitative RT-PCR to detect virus content of diseased plant. Results show that, the higher virus-carried in laodelphax striatellus, the more rich virus in plant, the earlier and the more serious disease occurred. When the virus-carried rate is23.3%, all the experimental maize material can be identified more certainty out of others. In this group, virus content between the maximum and minimum is largest.Levels of resistance of the various materials are:P138is moderate resistance, C7-2and Dan340is resistance, other materials are high susceptibility which virus-content order of magnitude is as high as106copy/μl. In the virus-carried rate of10%, some material content modicum amounts of virus, but have no incidence.The virus in disease-resistant materials have little proliferation, virus’s peak is significantly smaller than the high susceptibility of material. Susceptible materials following the linear multiplication of the virus in the body, and virus content have high peak.更多还原