Study on molecular mechanism of physiological specialization in Fusarium oxysporum

为研究尖孢镰刀菌的生理特化，建立了尖孢镰刀菌的转化体系，为融合细胞的选择提供耐药性依据。采用PEG法将携带耐湿霉素基因的质粒pHL1导入番茄尖孢菌原生质体中。通过对番茄植株的耐受性和致病性进行筛选，筛选出抗潮霉素b的菌落，并对R7-HygB进行转化。另一方面，用电法将黄瓜尖孢子菌直接转化为诺维信234处理后的孢子。番茄枯萎菌的毒力因子研究番茄枯萎菌产生多种果胶酶和镰刀酸作为毒力因子。果胶酶之一，聚半乳糖醛酸酶（PG），已被推测为一个重要的毒力剂。对真菌细胞进行诱变和非致病性筛选。获得的突变体OW932是一种非常弱的病原菌，与亲本相比PG蛋白含量较低，说明PG在尖孢镰秆的致病性中起重要作用。酶纯化后，SDS-PAGE和电泳分析显示有4种同工酶。进一步研究发现，用PG抗血清检测PG反应时，PG仅位于病变番茄组织的维管束上。因此，番茄枯萎菌引起的枯萎症状可能是真菌PG对维管束细胞的破坏。

Study on transformation and cell fusion of Fusarium oxysporumTo investigate on physiological specialization of F.oxysporum, effort was made to establish the transformation system of Fusarium fungus to confer drug resistance for selection of the fused cells. The plasmid pHL1 carrying hygromycinB-resistant gene was introduced into the fungal protoplasts of F.oxysporum f. sp. lycopersici by PEG method. The resuleted hygromycin B-resistant colonies were selected in degree of tolerance and in pathogenicity to tomato plants, and R7-HygB transformant was isolated. On the other hand, F.oxysporum f. sp.cucumerinum was transformed directly into the germed spores treated with Novozyme 234 by electropolation method.Study on virulence factor in wilt fungus of tomatoThe wilt fungus F.oxysporum f. sp. lycopersici produces various kinds of pectinases as well as fusaric acid as virulence factors. One of pectinases, polygalacuturonase (PG), has been speculated as an important agent for virulence. The fungal cells were mutagenized and selected in non-pathogenicity. The obtained mutant OW932 was shown to be a very weak pathogen This mutant was slight content in PG protein as compared with the parent strain, indication that PG plays an important role pathogenicity of F.oxysporum. After purification of the enzyme, SDS-PAGE and electrophoresis analyzes showed 4 kinds of isozymes. In further study, PG was located only at vascular bundle in diseased tomato tissues when PG reaction was detected using PG-antiserum. Thus, wilt symptom caused by F.oxysporum f. sp. lycoperisici may be damage of vascular bundle cells by the fungal PG.