【摘要】 以转座子mini Tn5为介导 ,用高压电激转化法将BtcryIA(c)基因整合到生防细菌荧光假单胞菌P30 3 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(c)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P30 3 1中的高效表达。生物测定结果显示工程菌对棉铃虫Heli coverpaarmigera具有显著毒杀作用。工程菌PT4 5、PT5 1、PT6 1、PT71和野生菌HD 73对棉铃虫 2龄幼虫 5天的LC50 值分别为5 0 12 81μg g、 71 776 3μg g、 6 9 0 82 0 μg g、 5 7 9895 μg g、 192 874 7μg g。死亡率与浓度呈正相关 ,体重与浓度呈负相关。更多还原

【Abstract】 Transposon mini Tn5 was used to integrate Bt cryIA(c) 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(c) 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 LC 50 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.更多还原