As surface-enhanced Raman scattering (SERS) is a convenient and sensitive method that can detect the analytes even on a single molecule level using a simple nanostructure, it has been employing as an attractive strategy for sensitive DNA sensing. Here, we report the development of a multiplex bacterial DNA detection method employing Au particleon-wire systems as a SERS sensing platform1. The Au NW-on-wire exhibits superb reproducibility, excellent sensitivity, and good time stability. A pattern formed by multiplex Au NW sensors provides positional address and identification for each sensor. By using this system, multiplex target DNAs were detected with a detection limit of 10 pM in a quantitative manner. The target DNAs obtained from reference strains and clinical samples were identified by the particle-on-wire SERS sensor developed in this study successfully, predicting practical values of this sensor in a variety of biosensing application where DNA binding involved.