大丽轮枝菌VdLac基因克隆与功能分析
Cloning and Functional Analysis of VdLac in Verticillium dahliae

为探讨漆酶基因VdLac在大丽轮枝菌中的功能，克隆该基因并利用农杆菌介导的遗传转化方法和PEG介导的原生质体转化方法对VdLac进行敲除和功能回复，获得敲除和互补突变体。以野生型JY为对照，对敲除突变体和互补突变体进行生物学功能测定。结果显示，VdLac基因全长1 978 bp，cDNA序列全长1 713 bp，编码570个氨基酸组成的蛋白质；VdLac基因缺失突变体( △VdLac)对硝化压力更敏感，其在CM培养基上仍可产生漆酶，但产生漆酶的能力降低；表明大丽轮枝菌漆酶基因VdLac不影响菌株的营养生长、产孢、致病力以及黑色素合成。
In order to explore the laccase gene(VdLac) function of Verticillium dahliae, the VdLac was cloned from the wild type strain JY of Verticillium dahliae.The deletion and its complementation were obtained through the gene knock-out and complementation methods by Agrobacterium tumifaciens-mediated transformation and PEG-mediated protoplasts transformation.Compared with the wild type strain JY of Verticillium dahliae, the deletion and complementation mutants were selected for VdLac biological function examination.The results showed that the complete DNA and cDNA sequences of the gene named VdLac were 1 978 bp and 1 713 bp in length, respectively, encoding a protein of 570 amino acid.The deletion mutants( △VdLac) were more sensitive to nitrosative stress, and could produce laccase on CM plates, but the production was reduced.The results indicated that VdLac had no effect on fungus vegetative growth, conidia production, pathogenicity and melanin synthesis in Verticillium dahliae.