辽宁省部分地区对鳞翅目害虫高毒力的苏云金芽胞杆菌菌株的分离与筛选  

Isolation and Screening of High Virulent Bacillus thuringiensis Strains Against Lepidopteran Pests in Some Areas of Liaoning Province

作　　者：倪鹤嘉 程雪 王静[1] 高泽平 魏盼盼 谭佳丽 孙敬 李海涛[1] 高继国[1] NI He-Jia;CHENG Xue;WANG Jing;GAO Ze-Ping;WEI Pan-Pan;TAN Jia-Li;SUN Jing;LI Hai-Tao;GAO Ji-Guo(College of Life Science, Northeast Agricultural University, Harbin 150030, China)

机构地区：[1]东北农业大学生命科学学院

出　　处：《农业生物技术学报》2019年第8期1513-1520,共8页Journal of Agricultural Biotechnology

基　　金：国家重点研发计划项目(No.2017YFD0201200)

摘　　要：鳞翅目害虫每年对农业生产造成巨大损失,化学农药的长期施用对生态环境造成严重破坏。苏云金芽胞杆菌(Bacillus thuringiensis, Bt)以其杀虫活性高,对环境友好等优点成为应用最为广泛的微生物杀虫剂。本研究拟对辽宁省苏云金芽胞杆菌的分布和基因多样性进行研究,了解其分布特点,发掘新型Bt资源,筛选出对鳞翅目害虫有高活性的菌株。利用温度筛选法从土壤中分离得到野生Bt菌株,利用限制性片段长度多态性PCR方法(restriction fragment length polymorphism PCR, PCR-RFLP)分析其基因型,利用SDS-PAGE对其杀虫晶体蛋白进行分析,并测定分离菌株对鳞翅目5种害虫的杀虫活性。从辽宁省不同地区收集不同类型的土壤524份,共分离出66株野生Bt菌株,出菌率为12.6%。晶体形状呈现出菱形、球形、短双锥、不规则形等形状。分离株多含有cry1、cry2及cry8基因,表达约130、90和60 k D的蛋白。杀虫毒力检测表明,菌株DD229、DD214-2、YK524和YK540对鳞翅目害虫具有较高活性。辽宁省地区苏云金芽胞杆菌分布广泛、资源丰富,获得4株对鳞翅目害虫杀虫活性较高菌株,具有良好的研究和应用价值。Lepidopteran pests cause huge losses to agricultural production every year, and long-term application of chemical pesticides cause serious damage to the ecological environment. Bacillus thuringiensis(Bt) is widely used as microbial insecticide because of its high insecticidal activity and environmental friendliness. The aim of this study was to isolate Bt strains from the soil of Liaoning province and to characterize them on the basis of the morphology of insecticidal crystal proteins(ICPs), the cry gene, the ICPs profiles and the insecticidal activities against lepidopteran pests. The temperature screening method was applied to isolate the wild Bt strains. The cry genotypes were analyzed by restriction fragment length polymorphism PCR(PCR-RFLP). The ICPs profiles were analyzed by SDS-PAGE, and the isolates were tested for their insecticidal activities against 5 species of Lepidopteran pests. A total of 524 soil samples were collected from different areas of Liaoning province, and 66 wild Bt strains were isolated, with a prevalence rate of 12.6%. The ICPs of the strains exhibited varied shapes such as rhombus, sphere, short biconical and irregular shape. The isolated strains expressed ICPs, including 130, 90 and 60 kD proteins. The insecticidal toxicity test showed that Bt strains DD229, DD214-2, YK524 and YK540 had high activities against the tested Lepidoptera pests. The results showed that Bt was abundant in the soils of Liaoning province. And 4 strains that showed high insecticidal activities against Lepidopteran pests which would have good research and application value.

关 键 词：苏云金芽胞杆菌(Bt) 限制性片段长度多态性PCR方法(PCR-RFLP) 鳞翅目害虫 CRY基因 

分 类 号：S436.3[农业科学—农业昆虫与害虫防治]