大豆疫霉菌的EMS化学诱变

以甲基磺酸乙酯(ethylmethane sulfonate,EMS)为诱变剂,通过其对大豆疫霉菌Phytophthora sojae休止孢萌发的影响,确定化学诱变条件。通过收集单卵孢子,建立了包含640个单卵孢子系的突变体库,其中约有50%的诱变菌系在培养性状和菌落形态方面发生了明显变化,菌落形态多样,表现出较紧密或松散,近圆形或不规则;气生菌丝减少,生长速度较慢或快;在卵孢子产量方面,8.13%的菌系有增加,20.41%的菌系减少,27.82%的菌系极少或者没有卵孢子产生,43.64%的菌系卵孢子产量类似野生型。以质膜氢离子泵蛋白基因PsPMA1(plasma membrane H+-ATPase1)为对象,通过TILLING技术,从320个大豆疫霉菌突变体中获得9个突变体,进一步确认了EMS对大豆疫霉菌的诱变效果,并且估算EMS对大豆疫霉菌的诱变频率至多每115kb发生一个核苷酸变异。新构建的突变体库为开展大豆疫霉病菌的功能基因组研究奠定了遗传材料基础。

EMS mutagenesis analysis of Phytophthora sojae

The commonly used chemical mutagen EMS (ethylmethane sulfonate) was employed to create a mutant collection by treating the encysted zoospores of Phytophthora sojae and the spore germination was used to optimize the mutagenesis condition. A total of 640 single-oospore mutant lines were acquired, among which 50% of them showed a wide range of morphological changes. As to the production of oospores, 8.13% mutants produced more, 20.41% less, and 27.82% very few or no oospores as compared with the wild type, respectively, and 43.64% were similar to the wild type. The quality of the mutant collection was further confirmed by successful identification of 9 mutations in PsPMA1 (Plasma Membrane H+-ATPase 1) gene from 320 P. sojae mutants. Thus, the estimated maximum frequency of EMS induced mutation is 1 per 115kb of sequence. The created P. sojae mutant collection will be useful for functional genomic analysis of P. sojae.