从老化病组织中高效分离大豆疫霉菌的方法  

A method for high-efficiency isolation of Phytophthora sojae from heavily infected soybean tissues

作　　者：宁峰[1,2] 朱晓莹[1] 殷丽华[1] 左豫虎[3] 单卫星[1,4] 

机构地区：[1]西北农林科技大学植物保护学院,陕西杨凌712100 [2]广东茂名市农业局,广东茂名525000 [3]黑龙江八一农垦大学植物科技学院,黑龙江大庆163319 [4]陕西省农业分子生物学重点实验室,陕西杨凌712100

出　　处：《西北农林科技大学学报（自然科学版）》2009年第5期139-143,共5页Journal of Northwest A&F University(Natural Science Edition)

基　　金：国家自然科学基金项目(30571204);教育部新世纪优秀人才支持计划项目(NCET-05-0856)

摘　　要：【目的】建立从老化病组织中高效分离大豆疫霉菌的方法,为大豆疫霉菌的群体遗传研究奠定基础。【方法】2006～2007年分别调查黑龙江和新疆大豆根腐病的发生情况,分别用病组织直接分离法和新建立的病组织浸泡分离法从采自黑龙江和新疆病样中分离大豆疫霉菌,统计分离率。【结果】建立的病组织浸泡法为:用NaOH处理采集的病样,通过镜检卵孢子的方法确定大豆疫霉病,排除镰刀菌、丝核菌或其他杂菌的干扰;浸泡病组织刺激卵孢子的萌发,用大豆叶碟诱集游动孢子侵染,下胚轴接种染病叶碟至感病大豆,最后从染病大豆胚轴分离病菌。采用此方法从采自黑龙江和新疆的205个已确认为大豆疫霉菌侵染的根腐病病样中,获得共计145个分离物,直接分离法和组织浸泡法平均分离率分别为21.4%和49.3%,总体分离率平均为70.7%。【结论】建立了从老化病组织中高效分离大豆疫霉菌的方法,降低了分离成本。[Objection] A method which could isloate Phytophthora sojae from heavily infected soybean tissues effectively was established,making it possible for large-scale isolation of P. sojae for population genetic analysis. [Method] From 2006 to 2007,investgation of root rot of soybean was made in Heilongjiang and Xinjiang individually. Directly isolated method and indirectly isolated method were used to isolate P. sojae from soybean samples respectively collected from Heilongjiang and Xinjiang, then the percentage of isolation was calculated. [Result] Indirectly isolated method ： NaOH treatment was used to exclude soybean root and stem rot caused by Fusarium, Pythium,Rhizoctonia and other pathogens by softening collected samples and checking for the presence of Phytophthora sojae oospores. The diseased soybean tissues confirmed to contain Phytophthora were soaked with soil extract and baited with susceptible soybean leaf. The infected soybean discs were hypocotyl-inoculated on susceptible soybean seedlings and the freshly diseased tissues were used to isolate P. sojae directly. The established method was successfully used to obtain 145 P. sojae isolates from 205 soybean samples collected from both Heilongjiang and Xinjiang that were confirmed to be infected with P. sojae. The average percentage of directly isolated method and indirectly isolated method was 21.4% and 49.3% respectively,and the avreage total percentage of isolation was 70.7%. [Conclusion] The newly established method made it possible to isolate P. sojae from heavily infected soybean tissues with high efficiency, and the cost of isolation was reduced.

关 键 词：大豆疫霉菌 卵孢子 叶碟诱集 病菌分离方法 

分 类 号：S435.651[农业科学—农业昆虫与害虫防治]