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枯草芽孢杆菌生防菌B-916对水稻纹枯病和稻曲病具有良好的防治效果,为拓宽其防治范围,兼治病虫害,本研究采用B-916内源芽孢形成期特异表达启动子成功实现了杀虫晶体蛋白基因Cry1Ac在野生型生防菌B-916的高效表达,成功构建了兼治水稻病、虫害的工程菌E-916。经SDS-PAGE电泳分析,异源Cry1Ac基因在对数生长后期其表达产物杀虫晶体蛋白占到胞内总蛋白的28%。杀虫和抑菌试验结果表明构建的工程菌对水稻二化螟具有杀虫活性,且保持了其原有对水稻纹枯病的拮抗性能。构建的工程菌具有良好的分离稳定性,连续培养25h,稳定性高于95%;连续培养50 h稳定性高于70%;连续稀释培养50 h,质粒丢失频率为每代小于10-3个。
Bacillus subtilis biocontrol bacteria B-916 on rice sheath blight and rice false smut has a good control effect, in order to broaden the scope of its prevention and control of pests and diseases, the study using B-916 endogenous sporulation of the specific expression of the promoter was successfully achieved The efficient expression of insecticidal crystal protein gene Cry1Ac in the wild-type biocontrol strain B-916 was successfully constructed, and the engineering strain E-916, which is also responsible for rice diseases and insect pests, has been successfully constructed. By SDS-PAGE analysis, the heterologous Cry1Ac gene expression insecticidal crystal protein accounted for 28% of the total intracellular protein in the late logarithmic growth phase. Insecticidal and bacteriostatic test results showed that the constructed engineering bacteria had insecticidal activity against rice stem borer, and maintained its original antagonism against rice sheath blight. The constructed engineering bacteria had good separation and stability, and the stability was higher than 95% after continuous culture for 25 hours. The stability was higher than 70% after 50 hours of continuous culture. The frequency of plasmid loss was less than 10-3 .