橡胶树炭疽菌候选效应蛋白基因Cg4LysM的分离鉴定和初步功能分析

发布时间：2018-06-14 00:06

本文选题：胶孢炭疽病菌 + Cg4LysM　；参考：《海南大学》2017年硕士论文

【摘要】：效应蛋白是病原真菌的主要致病因子之一。目前关于胶孢炭疽病菌LysM型效应蛋白的研究十分有限,本研究在前期研究的基础上克隆了橡胶树胶孢炭疽病菌的一个LysM型效应蛋白编码基因,命名为Cg4LysM,对其功能进行了初步的研究,得到以下结果:本研究通过RT-PCR技术扩增了Cg4LysM基因的cDNA全长序列,测序表明,该基因含1983个碱基对,编码一个660个氨基酸的多肽,由297个极性残基、208个疏水基残基、8个强碱性残基和59个强酸性残基。生物信息学分析结果显示,该蛋白含有4个保守的LysM结构域,,不含任何跨膜结构域,推测为胞外分泌蛋白。同源性比较分析结果发现,该蛋白与草莓胶孢炭疽菌(Colletotrichum gloeosporioides Nara gc5)、高粱炭疽病菌(Colletotrichum sublineola)、石膏样小孢子菌(Microsporum gypseum CBS)的LysM蛋白同源,同源性分别为97%、59%、55%。为了研究Cg4LysM基因的功能,我们通过同源重组的方法构建了Cg4LysM基因的敲除突变体(徐文晶等,2015;郭立佳等,2014;),命名为△Cg4LysM。对△Cg4LysM突变体的表型进行分析,得出以下结果:(1)与野生型菌株相比,△Cg4LysM的菌落形态和孢子形态没有明显变化,但菌落的生长速率降低,孢子产量下降;(2)△Cg4LysM对蔗糖的利用增强;(3)△CgLysM对橡胶树叶片的致病性减弱,但对寄主细胞的机械穿透力没有变化。以上结果表明,胶孢炭疽病菌Cg4LysM基因不仅参与病原真菌的生长发育和能量代谢过程,而且与病菌的致病性相关,但与真菌的机械穿透力无关。
[Abstract]:Effector protein is one of the main pathogenic factors of pathogenic fungi. At present, the research on LysM type effector protein of Gymnotrichum anthracnose is very limited. On the basis of previous studies, a LysM type effector protein encoding gene was cloned from Hevea gloeosporium anthracnose. Cg4LysM was named as Cg4LysM, and its function was preliminarily studied. The following results were obtained: the full-length cDNA sequence of Cg4LysM gene was amplified by RT-PCR. Sequencing showed that Cg4LysM gene contains 1983 base pairs and encodes a 660amino acid peptide. There are 297 polar residues, 208 hydrophobic residues, 8 strongly alkaline residues and 59 strongly acidic residues. Bioinformatics analysis showed that the protein contained four conserved LysM domains, without any transmembrane domain, and was presumed to be an extracellular secretory protein. The results of homology analysis showed that the protein was homologous to LysM protein of Colletotrichum gloeosporioides Nara, Colletotrichum sublineolaae, Microsporum gypseum CBSs of Colletotrichum sublineolaum and Microsporum gypseum CBSs, and the homology was 975955555.The protein was homologous to Colletotrichum gloeosporioides Nara Gc5, Colletotrichum sublineolaae and Microsporum gypseum CBS. In order to study the function of Cg4LysM gene, we constructed Cg4LysM knockout mutants (Xu Wenjing et al. 2015, Guo Lijia et al. 2014) and named Cg4LysM by homologous recombination. By analyzing the phenotype of Cg4LysM mutant, the following results were obtained: (1) compared with wild-type strain, Cg4LysM had no significant changes in colony morphology and spore morphology, but the growth rate of Cg4LysM decreased. The pathogenicity of CgLysM to the leaves of Hevea was weakened, but the mechanical penetration of host cells was not changed. The results showed that the Cg4LysM gene was not only involved in the growth, development and energy metabolism of pathogenic fungi, but also related to the pathogenicity of fungi, but not to the mechanical penetration of fungi.
【学位授予单位】：海南大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S763.7

【相似文献】