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目的:建立一种快速、准确鉴定环保微生物菌剂中常用5种芽孢杆菌的方法。方法:根据rpoB和gyrB基因序列分别设计扩增5种菌的特异性引物及Taqman探针,并进行反应条件优化。结果:5种芽孢杆菌均产生特异扩增信号,最低检出浓度为:枯草芽孢杆菌24 cfu/ml、地衣芽孢杆菌35 cfu/ml、巨大芽孢杆菌50 cfu/ml、短小芽孢杆菌41 cfu/ml、环状芽孢杆菌21 cfu/ml。结论:研究建立的实时荧光PCR鉴定方法具有良好的重复性和准确性,可用于微生物菌剂中5种常用芽孢杆菌的快速鉴定,具有很好的推广应用前景。
OBJECTIVE: To establish a rapid and accurate method for the identification of 5 Bacillus commonly used in environmental microbial inoculants. Methods: According to the sequences of rpoB and gyrB genes, five primers and Taqman probes were designed and synthesized respectively, and the reaction conditions were optimized. Results: The specific amplification signals of all five Bacillus species were detected. The minimum detectable concentrations were: Bacillus subtilis 24 cfu / ml, Bacillus licheniformis 35 cfu / ml, Bacillus megaterium 50 cfu / ml, Bacillus pumilus 41 cfu / ml , Bacillus circulans 21 cfu / ml. Conclusion: The established real-time fluorescent PCR identification method has good repeatability and accuracy, which can be used for the rapid identification of five commonly used Bacillus in microbial inoculum, and has good popularization and application prospects.