水杨酸途径中的防御基因在转草酸氧化酶基因油菜中的表达

采用RT-PCR方法检测对水杨酸依赖的信号途径中PR-1、GLU和CHI3个防御基因在转OXO基因与非转基因的油菜中表达差异的结果表明,在转基因油菜株系中这3个基因都有不同程度的上调表达,暗示转OXO基因的油菜抗菌核病的能力增强可能与对水杨酸依赖的信号途径受诱导有关。     

植病生防菌盾壳霉的研究进展

盾壳霉是世界性病原真菌核盘菌的主要拮抗真菌之一。许多研究表明盾壳霉具有控制温室和田间多种作物菌核病的潜力。目前欧洲一些国家已有盾壳霉商品制剂销售。为了加速盾壳霉产业化进程和更好地发挥其控制菌核病的作用 ,以及为了使人们对盾壳霉的研究不断深入 ,综述了盾壳霉生态学特性、遗传改良、对核盘菌的生防机制、在菌核病防治上的应用及基因工程研究等方面的研究进展。     

黄绿木霉菌代谢产物对大豆菌核病核盘菌的抑菌能力研究

利用室内实验的方法,研究了黄绿木霉对大豆菌核病菌的抑菌能力。黄绿木霉在与核盘菌（Sclerotinia sderotiorum）对峙培养过程中,5-7d即可将菌核病菌完全覆盖,在形成的菌核上会有黄绿木霉孢子出现,扫描电镜中观察到核盘菌的菌丝体发生变形;其代谢产物在经121℃处理25min后,抑菌作用仍可高达100％;在对黄绿木霉发酵过程中,利用PD培养基即可达到有效的控制大豆菌核病菌菌丝生长的目的,而且利用PD培养基发酵3-4d的黄绿木霉代谢产物抑菌效果最好,抑菌中浓度最低,发酵时间过长与过短,均不利于对病原菌的抑制。经黄绿木霉代谢产物处理的菌核菌电导率与呼吸强度均发生变化,在光学显微镜下观察到菌核菌菌体发生变化,且处理后的菌核菌致病力明显降低。     

利用SRAP分析核盘菌遗传多样性

核盘菌(Sclerotinia sclerotiorum)是危害油菜等多种经济作物的重要病原真菌.研究不同地区、相同或不同寄主核盘菌的遗传多样性对了解核盘菌的遗传演化过程和指导病害防控具有重要意义.我们采用序列相关扩增多态性(sequence-related amplified polymorphism,SRAP)标记对不同地理来源、不同寄主来源的76个核盘菌菌株的遗传多样性进行了分析.7对SRAP引物共获得260个位点,其中114个为多态位点,占43.85%.UPGMA聚类结果显示,在相似性系数为0.64时,76个核盘菌菌株分为4个组,每组包含的菌株数分别为54、18、2和2.聚类及主成分分析结果显示,来源于春油菜生态区和冬油菜生态区油菜上的核盘菌菌株可以明显分为两簇,而油菜、大豆、莴苣等不同寄主植物上的核盘菌菌株没有明显的遗传分化.分子变异分析(AMOVA)结果显示.不同地理来源、不同油菜生态区和不同寄主来源的核盘菌群体内的变异率分别为75.2%、81.2%和97.6%,均达到极显著水平(P<0.001);不同地理来源和不同油菜生态区的核盘菌群体间的变异率分别为24.8%和18.8%,也达到极显著水平(P<0.001);不同寄主来源的核盘菌群体间的变异率仅为2.4%,变异不显著(P=0.8673).研究结果表明,来源于春油菜生态区的核盘菌的遗传多样性高于冬油菜生态区.     

Physiological and Molecular Features of the Pathosystem Arabidopsis thaliana L.-Sclerotinia sclerotiorum Libert

The fungal pathogen Sclerotinia sclerotiorum Libert causes rot diseases on many crops worldwide and large economic losses occur frequently because of a lack of resistant varieties. The pathogenesis of S. sclerotiorum and the molecular basis of plant responses to the pathogen are poorly understood. In the present investigation, the process of S. sclerotiorum infection in Arabidopsis thaliana L., a plant that is highly susceptible to this fungus, was analysed. In addition, the defense activation in the host was investigated. A convenient inoculation method using millet grain was developed for S. sclerotiorum in Arabidopsis. The fungus rapidly infected the plants, probably through ball- or cushion-like infection structures. Visible symptoms developed within 24 h and plants were killed 72 h after inoculation. Cellulase, the main enzyme that caused host tissues to rot, was secreted by S. sclerotiorum in a pH-dependent manner. Oxalic acid, another pathogenic factor secreted by the fungus, induced necrotic lesions on the leaves, infection with S. sclerotiorum strongly induced the production of the pathogenesis-related （PR） proteins β-1,3-glucanase and chitinase in Arabidopsis. Furthermore, the PR gene PDF. 1 was induced, but not PR1, indicating that the pathogen activated basal defense of jasmonic acid/ethylene dependence, which is consistent with its necrotrophic characteristics. This pathosystem for Arabidopsis-S. sclerotiorum could provide an approach for the analysis of the interactions between S. sclerotiorum and other crops, thereby facilitating genetic manipulation techniques for controlling this pathogen.     

Ectopic expression of an expansin-like B gene from wild Arachis enhances tolerance to both abiotic and biotic stresses

Plant expansins are structural cell wall-loosening proteins implicated in several developmental processes and responses to environmental constraints and pathogen infection. To date, there is limited information about the biological function of expansins-like B (EXLBs), one of the smallest and less-studied subfamilies of plant expansins. In the present study, we conducted a functional analysis of the wild Arachis AdEXLB8 gene in transgenic tobacco (Nicotiana tabacum) plants to clarify its putative role in mediating defense responses to abiotic and biotic stresses. First, its cell wall localization was confirmed in plants expressing an AdEXLB8:eGFP fusion protein, while nanomechanical assays indicated cell wall reorganization and reassembly due to AdEXLB8 overexpression without compromising the phenotype. We further demonstrated that AdEXLB8 increased tolerance not only to isolated abiotic (drought) and biotic (Sclerotinia sclerotiorum and Meloidogyne incognita) stresses but also to their combination. The jasmonate and abscisic acid signaling pathways were clearly favored in transgenic plants, showing an activated antioxidative defense system. In addition to modifications in the biomechanical properties of the cell wall, we propose that AdEXLB8 overexpression interferes with phytohormone dynamics leading to a defense primed state, which culminates in plant defense responses against isolated and combined abiotic and biotic stresses.     

导入双价基因的转基因杂交油菜亲本及其对菌核病抗性的研究

比较了乙酰丁香酮、pH、共培养温度及不同激素配比对根癌农杆菌转化油菜(Brassica napus)的影响,建立了油菜高效转化体系.按该体系,将组成型表达几丁质酶和β-1,3-葡聚糖酶基因转化甘蓝型双低杂交油菜亲本恢复系和保持系,获得了转基因恢复系和保持系植株.对再生植株进行PCR和SouthemBlot检测,结果表明外源基因已整合到油菜基因组中.连续3代的PCR检测及转基因植株抗病性在自交后代中得到保持,证实外源基因已遗传到T3代.对转基因植株T1代离体叶进行菌核病菌丝体接种和T1及T2代大田自然侵染鉴定,结果表明,转基因恢复系棚40-12、棚40-32和保持系96B-2对菌核病的抗性比受体对照大幅度提高,大田鉴定其发病率连续2年均比受体对照减少78%以上,比抗病品种‘中油821'减少75%以上,病情指数比受体对照和‘中油821'减少均达显著水平,其抗病性能在后代稳定遗传,获得了高抗菌核病的转基因育种材料.     

Identification of prior candidate genes for Sclerotinia local resistance in Brassica napus using Arabidopsis cDNA microarray and Brassica-Arabidopsis comparative mapping

Arabidopsis was the leading model of dicot plant. An accomplished platform had been established for functional genomics studies. The platform had largely facilitated molecular biology research of Arabidopsis itself as well as research on its phylogenetic related plants[1,2]. Brassica napus, as an important cooking oil crop, had a close phylogenetic relationship with Arabidopsis[3―5]. In order to take advantage of avail-able Arabidopsis genetic and molecular tools, Girke et al. had explored …     

Factors Affecting Biological Control of Sclerotinia sclerotiorum by Fungal Antagonists

Studies were conducted to determine the effects of soil moisture (9, 16 or 24% w/w) and temperature (5, 15, 20 or 25°C) on the control of sclerotia of Sclerotinia sclerotiorum by five fungal agents in sterile and natural field soil. All five biocontrol agents were effective in reducing the survival of sclerotia of S. sclerotiorum in sterile soil under dry (9% moisture) or wet (24% moisture) conditions at 20°C, but only Coniothyrium minitans was effective in natural soil. Coniothyrium minitans was the most effective in reducing sclerotial viability at the temperature range of 15–25°C. Trichoderma virens was effective against sclerotia of S. sclerotiorum to a lesser extent than C. minitans , and in non-autoclaved soil, it performed best at 25°C. Although Epicoccum purpurascens , Talaromyces flavus and Trichothecium roseum were effective against sclerotia of S. sclerotiorum in some instances, they were less effective than C. minitans and T. virens . Sclerotia of S. sclerotiorum conditioned for myceliogenic germination were more vulnerable to attack by the biocontrol agents than dormant sclerotia. The implications are discussed with respect to enhancement of biological control of crop diseases caused by S. sclerotiorum in different geographic regions.