转基因球孢白僵菌发酵产物对野生菌株的增效作用  

Synergistic effect of fermented solution produced by transgenic Beauveria bassiana increasing the insecticidal efficacy of wild-type strain results from subtilisin-like activity

作　　者：张永军[1] 金凯[1] 罗志兵[1] 蒋小东[1] 裴炎[1] 

机构地区：[1]农业部生物技术与作物品质改良重点开放实验室

出　　处：《植物保护学报》2006年第3期246-250,共5页Journal of Plant Protection

基　　金：国家高技术研究发展计划"863"项目(2002AA245021);国家自然科学基金(30300235)

摘　　要：研究发现,超量表达类枯草杆菌蛋白酶基因CDEP-1的球孢白僵菌菌株Bb0062-15-4-CDEP-1的发酵产物对野生菌株具有明显的杀桃蚜增效作用。进一步分析增效作用与目的基因表达产物关系的结果显示,加入CDEP-1酶活为44.3±0.6U/μL的发酵产物,可使野生菌株对桃蚜的致死中浓度降低4倍以上,共毒系数(CTC)达404.6。但发酵产物经121℃处理20min,增效作用完全丧失。用饱和硫酸铵沉淀蛋白质分别将透析后的重溶蛋白质和上清液用于增效作用测定,结果证明增效成分主要是蛋白类物质。将梯度稀释的发酵产物与浓度为1×107孢子/mL的野生菌株孢子悬浮液混合进行生物测定,发现各处理的致死中时与CDEP-1的活性呈负相关,相关系数为-0.8946。由此推断,发酵产物中的增效物质主要是目的基因表达产物CDEP-1。We found that the ferment product of a transgenic Beauveria bassiana, overexpressing CDEP- 1, a cuticle degrading subtilisin-like gene, obviously increased the insecticidal efficacy of wild type strain against aphids Myzus persicae. To investigate the relationship between synergistic effect and CDEP-1 activity in the fermented solution,we mixed the ferment product （44.3±0.6 U/μL of CDEP-1 activity） with wild type conidia. Bioassay indicated that adding with the ferment product led to a more than 4-fold decrease in the median lethal concentration （LC50） of conidla against aphids, with cotoxicity coefficient （CTC） of 404.6. However, after the solution was heated at 121℃ for 20 min to quench the protease activity, the synergistic effect almost lost, revealing that the synergistic material in the fermented solution was thermosensitive. When the fermented solution was precipitated by saturated （NH4 ）2SO4, and then the pellets and the supernatant were dialyzed respectively, and bioassay showed that the synergistic effect existed mainly in the protein part. At the spore concentration of 1 × 10^7/mL, the median lethal time （LT50） of M. persicae was negatively correlated with CDEP-1 activity （the relative coefficient was -0. 8946）. The results suggested that the synergistic effect of the ferment product mostly resulted from subtilisin-like CDEP-1 activity.

关 键 词：类枯草杆菌蛋白酶 球孢白僵菌 发酵产物 

分 类 号：S476.1[农业科学—农业昆虫与害虫防治]