The R404A single machine two-stage compression refrigeration system for marine ultra-low temperature was simulated by EES when the evaporation temperature or the condensation temperature changes. Experimental verification of simulation results was carried out on the basis of the ultra-low temperature experimental platform consisting of R404A single machine two-stage compression refrigeration system with different conditions. The effect of evaporating temperature and condensing temperature on the performance of the single stage two-stage compression refrigeration system such as compressor exhaust temperature, COP and the compressor power was analyzed. The results indicated that with the decrease of evaporation temperature or increase of condensing temperature, the exhaust temperature of the compressor increased. The simulation results were in good agreement with the experimental results, the deviations value of the refrigeration performance between the experimental test and the simulation results was about 0.5, and the reliability of the model was verified. Experimental results showed that when the evaporating temperature rises or the condensation temperature drops by 5 ℃, the effect of evaporating temperature on COP was up to 7.8% than that of condensing temperature. The research results can provide reference for the follow-up work and have great practical engineering significance.