Two rapid and sensitive methods based on TaqMan qPCR and droplet digital PCR assay for quantitative detection of Bacillus subtilis in rhizosphere.

Aims Bacillus subtilis, a typical plant growth-promoting rhizobacteria, can benefit plant through promoting growth and reducing disease. The colonization intensity of B. subtilis in rhizosphere is a key factor for improving their effectiveness of field application. In this study, we developed a rapid and sensitive method for detecting B. subtilis in rhizosphere via TaqMan qPCR and droplet digital PCR (ddPCR) methods. Methods and Results The primers/probe set targeting gyrB gene could successfully distinguish B. subtilis from its close-related species. Both the TaqMan qPCR and ddPCR methods exhibited a good linear relationship in the sensitivity assay, suggesting the developed method was specific, effective and reliable. Finally, the two methods were used to detect the colonization dynamic of B. subtilis within Arabidopsis rhizosphere. Both of them showed a consistent trend compared with the traditional cultivation-based and microscopy-based methods. Conclusions The TaqMan qPCR and droplet digital PCR (ddPCR) methods we developed could be used to rapidly detect B. subtilis in rhizosphere. Significance and Impact of the study The TaqMan qPCR and ddPCR methods developed in this study can be applied to rapid quantitative detection of B. subtilis populations, and will be helpful to evaluate their effectiveness of field application.