Lactobacillus sobrius sp. nov., which was recently isolated from the intestine of weaning piglets, has potential probiotic properties. To follow the fate of L. sobrius strain 001T in dietary interventions, a novel and strain-specific quantitative detection procedure was developed. This procedure was based on the isolation of specific genomic fragments from the type strain by representational difference analysis and their detection by real-time PCR. The described strain-specific quantification approach may be used in studies aimed at tracking bacterial strains added to specific environments.