The major challenge of plastic electronics is the deposition of gold nanoparticles (AuNPs) on flexible substrates at room temperature. Here, we show fast, single step, room temperature deposition of AuNPs on polyethylene terephthalate (PET) and biaxially oriented PET (BoPET) substrate by employing dielectrophoresis. The deposition has been carried out using two-electrode system, with BoPET (or PET) and metallic (Pt or stain steel) mesh, under an AC signal of 20 kHz and 20 V peak-to-peak (V-PP) (signal for PET is 6 V-PP and 6 kHz). In this method, we show how to deposit AuNPs on PET-like insulator by exploiting its polarization capability under an AC signal. The polarization of PET has been confirmed by change in the Raman spectra of the PET film under in situ AC signals. Furthermore, we show that using this dielectrophoretic deposition method, the PET films can be patterned by AuNPs at room temperature without any pre- and posttreatment.