›› 2002, Vol. 45 ›› Issue (4): 419-424.

• 研究论文 •    下一篇


段灿星1, 张青文1, 徐静1, 张正伟1, 熊延坤1, 杨青芹2(1中国农业大学植物保护学院,北京100094; 2邯郸农业高等专科学校   

  • 出版日期:2002-08-20 发布日期:2002-08-20

Construction of insecticidal engineered strains of Pseudomonas fluorescens and their insecticidal activity

DUAN Can-Xing1, ZHANG Qing-Wen1, XU Jing1, ZHANG Zheng-Wei1, XIONG Yan-Kun1 , YANG Qing-Qin2   

  • Online:2002-08-20 Published:2002-08-20

摘要: 以转座子mini-Tn5为介导,用高压电激转化法将BtcryIA©基因整合到生防细菌荧光假单胞菌P303-1的染色体上。通过对重组菌株染色体DNA酶切分析、PCR检测,表明转化子DNA中含有cryIA©基因及其营养期表达的强启动子。SDS-PAGE鉴定、ELISA检测及电镜观察证实了杀虫晶体蛋白在P303-1中的高效表达。生物测定结果显示工程菌对棉铃虫Helicoverpa armigera具有显著毒杀作用。工程菌PT45、PT51、PT61、PT71和野生菌HD-73对棉铃虫2龄幼虫5天的LC50值分别为50.1281 μg/g、71.7763 μg/g、69.0820 μg/g、57.9895 μg/g、192.8747 μg/g。死亡率与浓度呈正相关,体重与浓度呈负相关。

关键词: 转座子mini-Tn5, 荧光假单胞菌, Bt cryIA©, 基因, 棉铃虫

Abstract: Transposon mini-Tn5 was used to integrate Bt cryIA© gene into the chromosome of the biocontrol bacterium Pseudomonas fluorescens P303-1 by electroporation. Digestion analysis and PCR amplification of the chromosomal DNA demonstrated the existence of the cryIA© gene and strong vegetative promoter in the chromosome of these transformants. SDS-PAGE,ELISA analysis and transmission electron microscope observation verified the efficient expression of insecticidal crystal protein in P303-1 Bioassay results showed these engineered bacteria were strikingly insecticidal to cotton bollworms. The LC50 of these bacteria against the second instar of cotton bollworms after five days was 50.1281 μg/g, 71.7763 μg/g, 69.0820 μg/g, 57.9895 μg/g, 192.8747 μg/g. Mortality of the treated larvae manifested a positive linear correlation with bacteria concentration,while weight showed a negative linear correlation with concentration.

Key words: transposon mini-Tn5, Pseudomonas fluorescens, Bt cryIA? gene, cotton bollworm