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马铃薯StBAG3基因的克隆及其在抗晚疫病过程中的功能研究

发布时间:2018-04-03 16:11

  本文选题:马铃薯 切入点:StBAG3 出处:《内蒙古农业大学》2017年硕士论文


【摘要】:BAG蛋白(Bcl-2 associated athanogene)家族是一类多功能蛋白家族,其功能广泛,在酵母、植物和动物中都存在。在动物细胞中,该蛋白既可调控细胞的凋亡,又参与肿瘤发生的过程。然而已有的研究表明,植物中的BAG蛋白介入植物的抗病防卫反应和逆境胁迫反应。本研究通过同源比对设计引物从马铃薯中克隆得到StBAG3基因,并成功构建了StBA4G的超表达载体和沉默载体。通过农杆菌介导的瞬时表达方法初步研究了StBAG3在马铃薯晚疫病抗性建立过程中的作用,具体的研究结果如下:1.通过分析拟南芥中已经克隆得到的AtBAG4的蛋白序列,找到BAG蛋白的功能域保守序列,以此序列在马铃薯基因库中比对得到StBAG3基因序列。以此序列设计引物,通过PCR克隆得到StBAG3基因。该基因全长2074bp,开放阅读框1026bp,编码341个氨基酸。蛋白质二级结构预测表明,该蛋白二级结构由α螺旋、β-折叠和无规则卷曲组成,其中α螺旋89个,β-折叠35个,无规则卷曲217个。蛋白结构的分析表明,StBAG3包含一个BAG功能域和一个UBQ功能域,没有跨膜结构,不产生信号肽,是一种亲水性的不稳定蛋白。蛋白亚细胞定位预测的结果显示StBAC3可能定位在细胞核。2.StBAG3在马铃薯中的表达谱分析结果表明不同马铃薯品种中的StBAG3表达量存在差异,其中冀张薯12号中的表达量最高,而夏坡蒂中StBAG3表达量最低,其他品种表达量居中。而StBAG3基因在马铃薯不同组织的相对表达量由高到低顺序是:茎、嫩叶、根和老叶。3.晚疫菌接种实验的结果表明,不同品种的马铃薯叶片中的StBAG3基因的表达量随着接种时间的推移而逐渐增加,并在48h达到峰值;接种72 h后,StBAG3基因的表达量开始下降。4.通过载体构建,获得了StBAG3在35S启动子驱动下的超表达载体和沉默载体。5.农杆菌介导的瞬时表达结果表明接种晚疫菌后瞬时表达StBAG3的马铃薯叶片的病斑扩展速率明显小于注射水和pCAMBIA1302-GFP空载体的叶片病斑扩展度。预示着StBAG3可能参与了马铃薯对晚疫病的抗性建立过程。
[Abstract]:BAG protein Bcl 2 associated DNA family is a kind of multifunctional protein family, which has a wide range of functions and is found in yeast, plants and animals.In animal cells, the protein not only regulates cell apoptosis, but also participates in tumorigenesis.However, previous studies have shown that BAG proteins in plants are involved in disease resistance, defense response and stress response.In this study, StBAG3 gene was cloned from potato by homologous alignment design primer, and the superexpression vector and silencing vector of StBA4G were successfully constructed.The role of StBAG3 in the establishment of resistance to late blight in potato was studied by Agrobacterium tumefaciens mediated transient expression method. The results are as follows: 1.By analyzing the protein sequence of AtBAG4 cloned from Arabidopsis thaliana, the conserved sequence of functional domain of BAG protein was found, and the StBAG3 gene sequence was obtained by comparing the sequence in potato gene bank.The primers were designed and the StBAG3 gene was cloned by PCR.The total length of the gene is 2074 BP, open reading frame 1026 BP, encoding 341 amino acids.The prediction of protein secondary structure showed that the protein secondary structure was composed of 伪 helix, 尾 -fold and irregular crimp. Among them, 伪 helix 89, 尾 -fold 35, irregular crimp 217.The analysis of protein structure shows that StBAG3 contains a BAG functional domain and a UBQ functional domain. It has no transmembrane structure and does not produce signal peptide. It is a hydrophilic unstable protein.The results of protein subcellular localization prediction showed that StBAC3 might be located in the nucleus. 2. The results of expression profiling of StBAG3 in potato showed that there were differences in StBAG3 expression in different potato varieties, and the highest expression level was found in Jizhanshu 12.However, the expression of StBAG3 was the lowest in Charlotte, while the expression of StBAG3 in other varieties was the middle.The relative expression of StBAG3 gene in different tissues of potato was in the order of stem, young leaf, root and old leaf.The results of inoculation experiment showed that the expression of StBAG3 gene in potato leaves increased with the inoculation time and reached its peak at 48h, and the expression of STBAG3 gene began to decrease at 72h after inoculation.The overexpression vector and silencing vector of StBAG3 driven by 35s promoter were obtained by vector construction.The results of transient expression mediated by Agrobacterium tumefaciens showed that the disease spot spreading rate of potato leaves with transient expression of StBAG3 after inoculation was significantly lower than that of water injection and pCAMBIA1302-GFP empty vector.It suggests that StBAG3 may be involved in the establishment of resistance to late blight in potato.
【学位授予单位】:内蒙古农业大学
【学位级别】:硕士
【学位授予年份】:2017
【分类号】:S435.32

【参考文献】

相关期刊论文 前10条

1 罗守进;;安徽省马铃薯主要病害的发生与防治[J];农业灾害研究;2015年10期

2 卢肖平;;马铃薯主粮化战略的意义、瓶颈与政策建议[J];华中农业大学学报(社会科学版);2015年03期

3 夏昒p,

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