抗锈病红小豆品种筛选及抗病机理研究

发布时间：2018-05-28 10:18

本文选题：红小豆 + 锈病　；参考：《黑龙江八一农垦大学》2017年硕士论文

【摘要】：红小豆锈病是红小豆生产中主要病害之一,严重影响红小豆的产量和品质。利用抗病品种是防治红小豆锈病最经济有效的措施,然而目前红小豆种质资源对锈病的抗性评价尚无研究报道,主栽品种均不抗锈病,生产上难有可利用的抗病资源,红小豆对锈病的抗病机制亦不明确。因此,本研究从红小豆资源对红小豆锈病的抗性评价入手,筛选抗病品种,研究红小豆锈病菌(Uromyces vignae)与不同抗性红小豆品种互作后,病原菌在叶片上侵染过程的差异,以及与抗病相关的CAT、SOD、POD、PPO活性和抗病相关基因的变化情况,为红小豆抗锈病资源利用奠定基础。研究取得主要结果如下:1.采用人工接种方法对85份红小豆资源进行抗锈性评价,获得2个免疫品种,16个高抗品种、10个中抗品种、26个中感品种和31个高感品种。抗病品种与感病品种相比,其对锈病抗性主要表现为接种锈病菌后显症及产孢的时间推迟2-3 d,并且产生夏孢子堆的密度小于与感病品种。2.利用荧光显微技术研究了锈病菌在不同抗性品种叶片上侵染过程的差异。锈病菌夏孢子在感病品种上于接种后6 h即可萌发并形成附着胞,24 h可产生气孔下囊和吸器母细胞,48 h形成胞内吸器,接种后8 d,叶片表面即可产生夏孢子堆。抗病品种与感病品种比较,夏孢子附着胞形成时间推迟,接种后12 h才形成附着胞,夏孢子萌发率和侵染率降低。在抗病品种中,接种后48 h,菌丝靠近的叶肉细胞出现抑制菌丝扩展的胞壁沉积物,产孢时间明显推迟。感病品种叶肉细胞不出现抑制菌丝扩展的组织结构。以上结果表明抗病品种对锈病菌的抗性表现为抗侵入和抗扩展两种特性。3.利用生理生化技术研究了与抗锈性相关的防御酶系活性的变化和抗病性的关系。结果表明:抗病和感病品种与红小豆锈病菌互作后,叶片中CAT、POD、SOD、PPO活性均有不同程度的升高。抗病品种和感病品种叶片中CAT和POD活性在接种后48 h达到第一个高峰,并且抗病品种中CAT和POD活性要高于感病品种,分别高30.2%和35.7%;抗病品种中,接种后12 h,叶片中的SOD活性比感病品种低25.9%,而接种48 h后,抗病品种中SOD活性要高于感病品种;抗病品种和感病品种中PPO活性在接种后12 h即开始升高,并且在整个互作过程中,抗病品种叶片中的PPO活性要始终高于感病品种。以上结果表明CAT、POD、SOD、PPO活性变化与品种抗病性呈正相关。4.利用qRT-PCR技术研究了几丁质酶(CHI)基因和β-1,3-葡聚糖酶(GLU)基因在不同抗性品种中的表达情况。结果表明:抗病品种接种红小豆锈病菌后24 h,CHI基因出现上调表达,并在接种后48 h表达量达到最大,此时相对表达量是对照处理的24.5倍。在感病品种中,CHI基因的表达量在接种后12 h开始上调表达,之后表达量开始下调,并与对照相比无明显变化;抗病品种中GLU基因相对表达量在接种后0-48 h内无明显变化,在接种后120 h明显上调,并且此时的表达量达到峰值,上调7.1倍。感病品种中GLU基因的相对表达量与抗病品种的变化趋势相似,但是感病品种中GLU基因的表达量上调幅度要低于抗病品种。以上结果说明CHI基因和GLU基因与品种抗锈性密切相关。
[Abstract]:Red bean rust is one of the main diseases in the production of red bean, which seriously affects the yield and quality of red bean. The use of resistant varieties is the most economical and effective measure to prevent red bean rust. However, there is no research report on the resistance evaluation of red bean germplasm resources to rust, and the main varieties are not resistant to rust, and it is difficult to make use of disease resistance in production. Resources, the resistance mechanism of red bean to rust is also not clear. Therefore, this study, starting with the evaluation of the resistance of red bean resources to red bean rust, screened the resistant varieties, and studied the difference of the infection process on the leaves of the pathogen of red bean rust fungus (Uromyces vignae) and the different resistant red bean varieties, as well as the CAT, S related to disease resistance. The changes of OD, POD, PPO activity and disease resistance related genes were used to lay the foundation for the utilization of red bean rust resistant resources. The main results were as follows: 1. using artificial inoculation method to evaluate the rust resistance of 85 red bean resources, obtained 2 immune varieties, 16 high resistant varieties, 10 medium resistant varieties, 26 medium varieties and 31 high sensitive varieties. Compared with susceptible varieties, their resistance to rust was mainly manifested by the delay of 2-3 d after the inoculation of rust fungus and the time of sporulation, and the density of the spore heap was less than that of the susceptible variety.2. using fluorescence microscopy to study the difference in the infection process of the rust fungus on the leaves of different resistant varieties. The spore of the rust fungus was susceptible to the disease. 6 h after inoculation can germinate and form attachments, 24 h can produce the inferior stoma and sucker mother cells, 48 h form the intracellular sucker, 8 d after inoculation, and the summer spore heap can be produced on the leaf surface. In the resistant varieties, 48 h after inoculation, the cell wall sediment which inhibited the mycelial expansion of the mycelium cells, the sporulation time was delayed obviously. The susceptible variety leaf cells did not have the tissue structure to inhibit the mycelium expansion. The results showed that the resistance of the resistant varieties to the rust fungus was two kinds of anti invasion and anti expansion. Sexual.3. uses physiological and biochemical techniques to study the relationship between the changes in the activity of defense enzymes related to rust resistance and the relationship between the resistance to disease. The results show that the activities of CAT, POD, SOD, and PPO in the leaves of the resistant and susceptible varieties and the red bean rust fungus increase in varying degrees. The activity of CAT and POD in the leaves of the resistant and susceptible varieties is 48 h after inoculation. The activity of CAT and POD in the resistant varieties was higher than that of susceptible varieties, 30.2% and 35.7%, respectively, and 12 h after inoculation, the SOD activity in leaves was 25.9% lower than that of susceptible varieties. After inoculation 48 h, the activity of SOD in the resistant varieties was higher than that of susceptible varieties, and the activity of PPO in the resistant and susceptible varieties was 12 h after inoculation. The PPO activity in the leaves of the disease resistant varieties was always higher than that of susceptible varieties during the whole process. The above results showed that the changes of CAT, POD, SOD, PPO activity and disease resistance were positively correlated.4. using qRT-PCR technology to study the table of the chitinase (CHI) gene and the beta -1,3- glucan enzyme (GLU) gene in different resistant varieties. The results showed that 24 h, CHI gene was up-regulated after inoculation of the red bean rust fungus, and the expression of 48 h reached the maximum after inoculation, and the relative expression was 24.5 times as high as that of the control treatment. In susceptible varieties, the expression of CHI gene was up to be expressed at 12 h after inoculation, and then the expression began to be downregulated and compared with the control. There was no obvious change in the relative expression of GLU gene in 0-48 h after inoculation, the 120 h increased obviously after inoculation, and the expression amount reached the peak and up to 7.1 times. The relative expression of the GLU gene in the susceptible variety was similar to that of the disease resistant varieties, but the expression of GLU gene in the susceptible varieties was expressed. The results showed that the CHI gene and GLU gene were closely related to the rust resistance of cultivars.
【学位授予单位】：黑龙江八一农垦大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S435.21

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