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分别选取了不同来源的三株松材线虫,对其可能的致病因子纤维素酶进行了基因克隆,其大小分别为1 159、1 138和1 159bp.随后的同源性分析结果表明这三个基因与已报道纤维素酶基因BX-eng-3具有较高的同源性,仅于含有较多重复序列处存在重复次数的差异,同时,由此获得的亲缘关系的结果与RAPD分析结果一致.对四种纤维素酶进行表达及活性确认表明四种纤维素酶水解纤维素能力相当,即纤维素酶所具有的致病性与酶活力自身无关,而是与其表达量相关,暗示存在差异的基因序列可能是纤维素酶的连接域,不影响酶的活性.选取其中一种纤维素酶BXC10进行松树纤维素水解实验,分别进行扫描电镜观察、还原糖分析和小角X射线衍射分析,结果均表明纤维素酶能有效水解松树纤维素,是重要的松材线虫病的致病因子,进一步为酶学学说提供了理论基础.
Three strains of Bursaphelenchus xylophilus from different sources were selected to clone their possible causative enzymes, which were 1 159, 1 138 and 1 159 bp, respectively.The subsequent homology analysis indicated that the three The results showed that there was a high homology between BX-eng-3 and BX-eng-3, which was only reported in the presence of more repeats. In the meantime, the results of genetic relationship and RAPD analysis The results showed that the four cellulases had the same capability of hydrolyzing cellulose, that is, the pathogenicity of cellulase had nothing to do with the enzyme activity itself, but rather its expression level, indicating the existence The differential gene sequence may be the connection domain of cellulase without affecting the activity of the enzyme.One of the cellulases BXC10 was selected to perform pine cellulose hydrolysis experiments, and the scanning electron microscopy, the reducing sugar analysis and the small-angle X-ray diffraction analysis were performed respectively, The results showed that cellulase could effectively hydrolyze pine cellulose, which is an important virulence factor of pine wood nematode disease, which further provided the theoretical basis for the enzyme theory.