Plant growth-promoting rhizobacteria (PGPR) have been reported to influence plant growth, yield, and nutrient uptake by an array of mechanisms. Uncovering the behavioral dynamics of PGPR is one of the most important issues necessary for understanding their functional performances. In this study, strain NJAU-Z9 which was found to possess complex functions and efficient rhizospheric colonization ability was selected from plenty of bacterial strains isolated randomly from the pepper rhizosphere soil and identified as Bacillus velezensis. Repeated seedling nursing tests performed absolute growth-promoting advantage for the novel isolated strain. After that, primers for the quantitative detection were designed based on its whole genome sequence (WGS), and a real-time PCR method was utilized to explore strategies for monitoring the strain in natural soil and in the pepper rhizosphere. Results showed based on the whole genome, two primers were identified as NJAU-Z9-specific quantitative PCR primers. Two seasonal pot experiments demonstrated that strain NJAU-Z9 effectively colonized the rhizosphere measured by the novel abundance detecting strategy, improved plant growth, and showed a positive correlation between bacterial number and biomass. This study offers a strategy based on a real-time PCR method for directly monitoring B. velezensis strain NJAU-Z9 in the soil and the rhizosphere and provides a reference for the quantitative study of other PGPR strains based on WGSs.