The polymerase chain reaction (PCR) chips of different metal film thicknesses were fabricated, and the PCR machine with/without a top heater and the single-side/double-side heater PCR chips were compared. Before fabricating the chips, one simulated the effect of various thin-film thicknesses and the temperature distributions of various powers, and it helped one to find the optimum parameters. The total substrate material was cyclic olefin copolymers, and the gold film was fabricated on the substrate by evaporation. The results after PCR for the deoxyri bonucleic acid (DNA) of an Escherichia coli TG-1 cell were obtained. The results of the DNA amplifying were , and these were 78–85% of the result of PCR machine with a top heater. There was no PCR product in the PCR machine without a top heater and single-side PCR chip. The thermal deformation temperature, thermal conductivity, and substrate thickness are the important parameters in a polymer substrate PCR chip, and the thermal deformation temperature is the key parameter.