Thermocline layer have remarkable effects on acoustic propagation in Persian Gulf environment. So far, no comprehensive research has been conducted to explore thermocline layer, especially its characteristics including top, thickness, and thermal gradient of thermocline in Persian Gulf. Besides, effects of thermocline on underwater acoustic propagation including transmission loss and sound channel formation in Persian Gulf have not been investigated as most studies to date have focused on the effect of thermocline on transmission loss in low frequencies, while high frequencies have been studied less. Accordingly, this study attempts to shed light on these ambiguities. ROPMI Cruise measurement data collected during summer (August) of 2001 in Persian Gulf were used in this study. To simulate the acoustic propagation, ray-tracing theory was also applied to make possible an analysis of high frequencies. Results showed that presence of thermocline layer have remarkable effects on transmission loss. Increasing the gradient in thermocline layer will raise transmission loss.