玉米大斑病单基因抗性与超氧物歧化酶的关系

 1989~1991年,以两套玉米抗、感大斑病近等基因系为材料,在同一遗传背景中分析比较了Ht1、Ht2、HtN等抗病和ht感病基因型材料在感染玉米大斑病菌(Exserohilum turcicum(Pass) Leonard and Suggs)过程中超氧物歧代酶(SOD)活性及其同工酶的动态变化.大量分析资料表明,与感病材料比较,各种抗病材料在病原物浸染初期SOD活性下降,酶活性较低.     

玉米灰斑病抗性机制中活性氧代谢的作用

研究了玉米灰斑病菌侵染四个抗病和感病的玉米品种时，叶片内部活性氧代谢酶及细胞过氧化产物含量的动力学变化。结果表明，抗、感病品种的SOD、CAT及POD酶活性在病茵侵染后都变化显著，抗病品种各酶活性变化幅度比感病品种大。叶片内过氧化产物MDA含量则相反，抗病品种沈试29在接种第13天时叶片MDA含量只为18．46nmol／g，而感病品种铁单9为23．14nmol／g，抗病品种比感病品种增加幅度小。说明活性氧代谢在植物抗病机制中起着重要作用，抗病品种对活性氧代谢的酶调节能力强，病菌侵染后细胞过氧化程度低，在痛菌侵染时活性氧清除酶活性最大增加值与发病程度呈显著正相关。     

串珠镰刀菌引起玉米穗粒腐病防御酶变化及其电镜观察

 对串珠镰刀菌(Fusarium moniliforme)侵染引起玉米穗粒腐病的防御酶活性变化和病原菌侵染过程进行研究。采用人工接菌的方法,分别对抗(Bt鄄1)、感(掖478)玉米材料进行接种,取抗、感材料间隔24 h 的6 个时间段接菌部位的苞叶组织,分析玉米植株感病后部分防御酶、同工酶谱的动态变化,并用扫描电镜对病原菌入侵植株过程进行组织病理学观察。扫描电镜观察发现,菌丝首先要经过1 ~ 3 d 生长后,大约在72 h 左右开始侵入气孔,并且随着时间的推移,侵入气孔的菌丝量逐渐增多。这说明病原菌是直接通过气孔侵入寄主苞叶组织。同时,玉米受串珠镰刀菌侵染后,苯丙氨酸解氨酶 (PAL)和过氧化物酶(POD)的活性都是先上升后下降,在感病材料Ye478 中PAL 的活性要比抗病材料Bt鄄1 中增加的更快、更高;同样对于POD 来说,在感病材料Ye478 中的活性要比抗病材料Bt鄄1 中的高,但变化趋势在2 个材料中相似;而丙二醛(MDA)的含量则相反,在感病材料Ye478 中的活性要比抗病材料Bt鄄1 中的低;对POD 同工酶酶谱分析,2 个材料都增加了3 ~ 4 个条带,没有明显的区别,这说明玉米感病后会通过增加POD 的活性来抵御外源病菌的侵入。总体而言PAL和POD 活性水平与材料抗性呈负相关;MDA 与材料抗性呈正相关关系。对玉米植株感病后防御酶活性变化的分析和病原菌入侵寄主的电镜观察结果,可为深入研究玉米穗粒腐病抗病机制和抗病育种提供参考。     

环境与寄主条件对玉米灰斑病菌侵染寄主的影响

利用玉米灰斑病菌株20-47、感病玉米品种(掖单13)和抗病玉米品种(沈单10号),在人工气候室和自然条件下对影响玉米灰斑病菌侵染的环境条件和寄主生育期进行了研究。结果表明:玉米灰斑病是属偏高温高湿类型的病害,在温湿度条件不能得到满足时,病害就难以完成侵染发病;病菌接种侵入的最佳温度为25℃左右,水滴条件下侵染最容易;土壤条件特别是氮、磷肥如果施用不均匀、不足或过多,也都可以对结果产生影响,在通常情况下,增施氮、磷肥能提高玉米对灰斑病的抗性;在一定光照强度范围内,光对鉴定结果的影响不明显,但光暗交替更有利于寄主发病;寄主不同生育期对病原菌侵染寄主的影响不同,11～12叶期(喇叭口期)接种,病菌较易侵染。     

玉米圆斑病防治研究

目前吉林省推广的玉米骨干自交系吉63果穗上发生的穗腐症状是由Hel-minthosporium carbonum Ullstrup侵染引起的圆斑病。从病斑形态和培养性状与小斑病菌迥然不同。经温室苗期和田间成株期人工接种证明,圆斑病菌除为害吉63和少数自交系外,不侵染T型、M型等不育系及多数正常系玉米的果穗。从果穗抽出到灌浆期是圆斑病菌侵染发病的关键时期。病菌主要在病株残体里以菌丝体越冬,已腐烂的病组织则丧失生活力。六年共鉴定655份自交系、品种及杂交种,有184份材料的果穗感病,以吉63及其选系后代发病最重。经测定吉63感染圆斑病是由隐性感病基因所控制。     

利用mRNA差异显示技术分离和克隆玉米抗大斑病相关基因片段

 玉米大斑病是由玉米大斑病菌(Setosphaeria turcica)引起的玉米叶部主要病害。在玉米的抗病反应中玉米与大斑病菌间存在着复杂的互作关系。要阐明玉米抗病过程中抗病基因、参与信号转导基因、防卫反应基因等如何参与抗病过程,以及它们与抗病性的关系,需从基因表达整体水平上了解上述抗病相关基因表达的种类与数量,进而分析其相互关系。     

玉米灰斑病抗性反应中酚类物质代谢作用的研究

 研究了玉米灰斑病菌(Cercospora zeae-maydis)侵染前后的4个玉米品种苯丙氨酸解氨酶、过氧化物酶、多酚氧化酶及木质素含量的变化。3种酶比活性在病菌侵染后都发生明显的先增后降的变化,而抗病品种的变化要明显大于感病品种的变化,尤其是苯丙氨酸解氨酶最大增加活性与品种抗病性呈现极显著的相关性。4个品种的木质素含量在病菌侵染的第9 d时增加到最大值,以后略下降,而且抗病品种的木质素含量峰值高于感病品种的峰值。     

细胞微丝骨架在玉米抗纹枯病菌Rhizoctonia solani 侵染中的作用

为探究细胞微丝骨架在玉米抗纹枯病侵染过程中的作用,采用微丝骨架解聚剂LatB预处理玉米离体叶片后接种立枯丝核菌Rhizoctonia solani AG1-IA,显微观察病原菌的侵染过程,并检测活性氧(ROS)、细胞坏死及抗病基因(PR1、ZmDREB2A)表达等抗病反应情况。结果显示,与未经LatB预处理相比,LatB预处理加快了R.solani侵染后玉米病斑的形成,并影响了侵染结构的发育;在侵染后期,LatB促进了R.solani诱导的玉米叶片中ROS积累、细胞坏死反应和PR1基因表达;溶剂DMSO预处理与未经LatB预处理的结果类似,表明DMSO对本试验的影响较小。研究表明,细胞微丝骨架不仅参与玉米抗R.solani的侵入,而且通过调控ROS、PR1基因表达,细胞死亡等抗病信号提高玉米抗病防御能力。本研究为进一步研究细胞微丝骨架在玉米对纹枯病的抗病机理中的作用提供了重要参考。     

玉米灰斑病菌的可溶性蛋白质及同工酶多态性

利用聚丙烯酰胺凝胶电泳技术,对采自北方玉米主产区的23个玉米灰斑病菌菌株进行可溶性蛋白质和同工酶电泳图谱分析及聚类分析,从蛋白质和酶学的多态性水平上分析玉米灰斑病菌的生理分化特征.研究表明,玉米灰斑病菌在可溶性蛋白质和SOD、MDH、PPO、POD、EST、CAT等的同工酶谱存在差异,不同菌株之间某些同工酶谱带数和同一迁移率谱带的亮度和色泽差异非常显著,说明菌株间的多态性可在同工酶水平上得到反映.研究还发现,来自不同地区的菌株同工酶谱带无明显的变化规律,反映出病菌同工酶的变异与地理位置关系不密切,也表明该病菌可能具有较广泛的地域适应性.     

Ht2背景下玉米对大斑病菌1号小种抗性基因的表达差异研究

以玉米自交系黄早四和黄早四Ht2近等基因型系为材料,利用cDNA-AFLP差异显示方法,分析玉米抗大斑病Ht2相关基因受大斑病菌1号小种胁迫后的差异表达情况。用筛选的70对引物在黄早四Ht2中扩增出76个差异显示片段,对差异条带回收、克隆和测序,并在NCBI上进行BLAST分析,有52个在GenBank中发现相似性较高、功能已知的EST序列,按其功能分为以下8类:基础能量代谢、跨膜运输蛋白、抗逆或抗病相关蛋白、蛋白质代谢、染色体组成蛋白及DNA复制和转录中的蛋白、信号传导、生长发育调控因子和转录因子,而其中以参与基础能量代谢的基因和抗逆或抗病相关基因所占比例较大。功能已知的52个差异表达片段在玉米1～9号染色体上均有分布,且在非亲和互作前期(6h)主要是一些信号相关基因的表达,12h表达的基因多属于抗病反应初期阶段的相关基因,与防卫有关的基因主要在48～72h表达,生长发育类基因在所检测的非亲和互作的不同阶段均表现作用,在非亲和互作后期主要是蛋白质代谢基因的表达。     

东北春玉米区大斑病菌生理小种鉴定

对2005年采自东北玉米主产区的大斑病叶标样进行分离,经单孢纯化后获得56个菌株。通过Ht单基因鉴别寄主接种鉴定,结果表明:东北地区玉米大斑病菌生理分化已呈现复杂化,0号生理小种和1号生理小种分别占35.7%和17.9%,为优势小种,其次为123号生理小种,占12.5%,另外还存在2、3、23、123N、12和23N号生理小种。     

Physiological races ofExserohilum turcicum in Israel

A set of differentials of corn plants(Zea mays L.) containing Ht1, Ht2, Ht3 or HtN genes was used to identify races ofExserohilum turcicum in Israel. Plants were inoculated with 14 isolates ofE. turcicum collected from various regions in Israel (from Ayyelet HaShahar in the north to Sa’ad in the south). Differentials containing Ht1, Ht2, Ht3 or HtN genes were resistant to the 14 isolates tested, whereas the inbred lines without Ht genes were highly sensitive. Resistance was characterized by the formation of non-sporulating chlorotic lesions. When plants containing Ht1, Ht2 or Ht3 genes were inoculated with relatively high inoculum concentrations (over 50 conidia/drop), chlorotic lesions were associated with necrosis in the center of the lesions. Sporulation of the fungus in the necrotic parts of the lesions was significantly less than on plants without Ht genes. No necrosis was observed in plants with the HtN gene. Our results indicate that the physiological race ofE. turcicum in Israel is race 1.     

玉米大斑病菌毒素结构的确定及几种类似物的毒性比较

 通过对玉米大斑病菌Ht-毒素组分1与标准5-羟甲基-2-呋喃甲醛核磁共振¹H谱比较,进一步明确了组分1的化学结构。用玉米离体叶浸渍法及离体根冠细胞致死生测法对Ht-毒素组分1、5-羟甲基-2-呋喃甲醛、呋喃甲醇、呋喃甲醛、呋喃甲酸进行毒性和致病活性比较,结果表明,Ht-毒素组1、标准样品及几种毒素类似物在供试浓度范围内对寄主玉米均有高度的生物毒性,其中以呋喃甲酸毒性最大。从TLC扫描光谱和色谱来看,5-羟甲基-2-呋喃甲醛确为Ht-毒素的一种主要毒性组分,而且该化合物在体内外随着不断氧化,生物毒性逐渐提高。     

中国玉米大斑病菌生理分化及新命名法的初步研究

 在分别含Ht1、Ht2、Ht3和HtN的单基因玉米鉴别寄主上测定了全国玉米各产区的百余份大斑病菌标样。结果表明,按传统方法命名的1号(或称0号)生理小种仍然是优势小种,占供试菌株的73.0%;2号(1号)生理小种占1.9%。虽然未出现已报道的3号(23号)、4号(23N号)和5号(2N号)小种,但特别值得注意的是出现了25.1%的未定名的新类群。为克服传统命名方法的局限性及指导抗病育种及品种布局,本文初步提出用毒力频率法分析玉米大斑病菌的生理分化状况,并进一步指明了不同致病类群在我国的分布频率及Ht基因的可用范围。     

Frequency of the Ht1 Gene in Populations of Sweet Corn Selected for Resistance to Exserohilum turcicum Race 1

ABSTRACT The possibility that the Ht1 gene or genes tightly linked to Ht1 convey general resistance to races of Exserohilum turcicum that are virulent against Ht1 (i.e., residual resistance) could be useful in sweet corn where the Ht1 gene is present in many commercial hybrids and breeding populations. The objective of this study was to determine if the frequency of the Ht1 gene changed in populations of sweet corn selected for general resistance to E. turcicum race 1, thus conveying residual resistance. Four populations were developed with theoretical initial frequencies of the Ht1 gene of 0, 0.25, 0.25, and 0.5. The populations were advanced by recurrent mass selection with parental control through four or five cycles of selection following inoculation with an Ht-virulent race of E. turcicum (i.e., race 1). Plants from each cycle of each population were evaluated for severity of northern corn leaf blight (NCLB) and chlorotic lesion reactions following inoculation in field and greenhouse trials with either race 0 or 1 of E. turcicum. Recurrent mass selection for general resistance to E. turcicum race 1 reduced the severity of NCLB in all four populations of sweet corn, although the change in the most susceptible population was minimal. Percent gain per cycle was 14.5, 12.3, 14, and 3.7% for populations I, II, III, and IV, respectively. The Ht1 gene did not convey levels of general resistance to E. turcicum race 1 that were substantial enough to be selected for in this population improvement program. There was no apparent selection advantage for resistance to E. turcicum race 1 in the populations that contained the Ht1 gene. The frequency of the Ht1 gene did not differ among cycles of selection within any of the populations in response to improved levels of general resistance to NCLB. The lack of change in frequency of Ht1 in these populations and the similarity in gain per cycle among populations with and without Ht1 lead us to conclude that the Ht1 gene itself did not have a residual effect on resistance.     

玉米抗大斑病HtP单基因鉴别寄主SSR标记的筛选

应用SSR标记技术,以含有Ht单基因的玉米自交系黄早四为试材,选用分别位于玉米10条染色体上的121对SSR引物经PCR扩增检测分析,筛选到黄早四Ht1基因有效标记9对,Ht2基因有效标记6对,Ht3基因有效标记1对,HtN基因未筛选到有效标记。     

玉米大斑病菌毒素对玉米叶肉细胞脂氧合酶活性的影响

 玉米大斑病菌2号小种粗毒素(HT-C)和标准毒素(5-羟甲基-2-呋喃甲醛,HT-I)处理玉米叶片,都能诱导3种供试基因型玉米叶片中脂氧合酶活性增高,但是不同基因型玉米对HT-C和HT-I的反应不同,HT-I对含有Ht1基因的品系诱导时间早,水平高,HT-C对于不带Ht基因的品系LOX的影响略高于HT-I;不同浓度的HT-I在OH43和OH43Ht1玉米叶片中诱导的LOX酶活性不同,高浓度HT-I和HT-C对LOX酶活诱导时间早,水平高,浓度降低诱导水平降低;外源H₂O₂对HT-I和HT-C在OH43叶片中LOX酶活性诱导作用的影响不同。     

玉米杂交种和自交系对多种病害抗病性的诱发病圃鉴定

在华农实验农场设置病圃,分区块重复分别接种大、小斑病、茎腐病、纹枯病、褐斑病和自然发生斑点病。从1980～1982年鉴定生产上推广的玉米杂交种和其亲本,以及培育中的材料共260份。其中多数抗3～4种病害;少数不抗到抗1～2病害,以至抗5～6种。湖北省低山地区推广三系杂交种“唐五”中抗到抗大、小斑病、茎腐病、褐斑病、斑点病,感纹枯病;二高山地区推广单交种“恩单2号”中抗大斑,中感小斑,中抗到中感茎腐、纹枯,中抗到抗褐斑和斑点。在培育中的新自交系“唐Mo17Ht_1cms-S”和“77cms-C”中抗到抗6种病害。根据国内外资料分析了抗小斑病材料“Mol7”含有加性显性抗病基因,并兼抗褐斑病;抗大斑病材料中包含Ht类加性显性抗病基因,以及抗茎腐病的多抗基因。又根据调查,讨论了进一步培育多抗材料,结合宽窄行套种、改夏播为春播、增施氮磷钾肥料、培土作垅等耕作栽培制度,保持和提高玉米品种的多种病害抗病性。     

Temporal Variation in Setosphaeria turcica Between 1974 and 1994 and Origin of Races 1, 23, and 23N in the United States

ABSTRACT Setosphaeria turcica causes northern leaf blight, an economically important disease of maize throughout the world. Survey collections of S. turcica isolates from 1974 to 1994 provided a unique opportunity to examine temporal diversity in the eastern United States. Two hundred forty-two isolates of S. turcica from maize were studied with random amplified polymorphic DNA (RAPD) markers, mating type, and virulence on maize differential inbred lines with known Ht resistance genes to examine changes over time. One hundred forty-nine RAPD haplotypes were identified. Nearly 20% of haplotypes recurred in more than one year. Race 0 isolates declined in frequency from 83% in 1974 to near 50% in the 1990s, most likely in response to the widespread deployment of Ht1 in commercial maize hybrids. Races 23 and 23N were present in the collection at low levels throughout the study period and were also found among isolates from Virginia in 1957. The frequency of MAT1-2 isolates increased sharply after 1979 and was associated with the emergence of race 1 during the same period. RAPD markers were used to investigate the genetic diversity among a subset of isolates collected in the United States from 1976 to 1982, the period in which this dramatic shift in race frequency occurred. Multilocus haplotypes were not exclusively associated with known races of S. turcica. Based on shared haplotypes and cluster analysis, race 1 isolates share greater similarity with race 0 than with 23 or 23N isolates, indicating race 1 probably evolved from multiple lineages of race 0. Sorghum spp.-infecting isolates share greater similarity with one another than with maize-infecting isolates and represent a distinct subgroup.