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The thermal cycle system is a key module for the amplification of deoxyribonucleic acid in real-time polymerase chain reaction (PCR) instrument. The accuracy of quantification is directly influenced by the performance of the system, such as heating and cooling rate, temperature uniformity and temperature-controlled precision. Therefore, a proper thermal analysis method is very important to the thermal cycle system design. In this paper, an equivalent circuit model of the system using thermoelectric coolers was proposed. The modeling results were in good agreement with the experimental results. The use of this model allowed the static and dynamic performance of the system to be analyzed by using electrical circuit analysis software such as Multisim10. Also, it is an effective and simple way to study how the factors such as thermal resistance of heat sink and thermal grease affect the performance of the system.