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A readout approach for the Hamiltonian path problem (HPP) in DNA computing based on the real-time polymerase chain reaction (PCR) is re-implemented on DNA Engine Opticon 2 System. Several types of fluorescent probes and detection mechanisms are currently employed in real-time PCR, including SYBR Green, molecular beacons, and hybridization probes. Based on the new approach, real-time amplification performed using the TaqMan probes is adopted, as the TaqMan detection mechanism can be exploited for the design and development of the proposed readout approach. In this study, double-stranded DNA molecules of length 140 base- pairs are selected as the input molecules, which represent the solving path for an HPP instance. These input molecules are prepared via the self-assembly of 20-mer and 30-mer single- stranded DNAs, by parallel overlap assembly. The proposed readout approach consists of two steps: real-time amplification in vitro using TaqMan-based real-time PCR, followed by information processing in silico to assess the results of real-time amplification, which in turn, enables extraction of the Hamiltonian path. The experimental result is compared with that of previously implementation on Roche LightCycler System. Experimental results establish an easier method to interpret the output of real-time PCR for the subsequent in silico information processing.