ABSTRACT The detection of soil-borne pathogens by quantitative PCR (qPCR) has been challenging due to the pronounced influence of soil type on DNA extraction and PCR reactions. In the present study, we developed a novel qPCR system and an internal sample process control (ISPC) strain, RsPC, for the detection of Ralstonia solanacearum species complex (RSSC), the pathogens causing bacterial wilt. Specific primers and TaqMan probe were designed based on analyses of 16S rRNA gene sequences from 581 Ralstonia genomes, and the RsPC was constructed by insertion of an artificial fragment, which consisted of two fragments from the kanamycin-resistant gene and the gfp gene, into the chromosome of a phylogenetically closely related strain, Ralstonia pickettii JCM 5969. The qPCR target sequences of RSSC and RsPC shared primers; however, different TaqMan probes were used to distinguish them from each other. The interaction assay between ISPC and target DNA showed no influence on sensitivity when their difference in concentration was