大豆疫霉菌的EMS化学诱变-单卫星试验室.PDF

Mycosystema 菌 物 学 报 15 March 2010, 29(2): 222-227 jwxt@ ISSN1672-6472 CN11-5180Q ©2010 Institute of Microbiology, CAS, all rights reserved. 大豆疫霉菌的 EMS 化学诱变 * 哈霞 胡中慧 王蕾 权军利 单卫星 西北农林科技大学植物保护学院 陕西省农业分子生物学重点实验室 杨凌 712100 摘 要：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+-ATPase 1 ）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。 关键词：卵菌，卵孢子，化学诱变，突变体库 EMS mutagenesis analysis of Phytophthora sojae * HA Xia HU Zhong-Hui WANG Lei QUAN Jun-Li SHAN Wei-Xing College of Plant Protection and Shaanxi Key Laboratory of Molecular Biology for Agriculture, Northwest Agriculture Forestry University, Yangling 712100, China Abstract: 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 mutati