微生物菌剂中5种芽孢杆菌实时荧光PCR鉴定

Identification of five Bacillus species used in environment microbe agentia with real-time PCR

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 to identify the five Bacillus species generally used in environment microbe agentia.Methods：By analyzing nucleotide sequences of rpoB and gyrB genes,the species-specific primers and probes were designed and the reaction conditions were optimized in this research.Results：The result showed that all five species generated a specific amplification signal.The detection limit of B.subtilis,B.licheniformis,B.megaterium,B.pumilus and B.circulans was 24,35,50,41 and 21 cfu/ml,respectively.Conclusion：The specific real-time PCR method established in this study has good repeatability and accuracy,and has very good prospects for the application in rapid identification of these Bacillus species.