The heat treatment process of 34CrMo4 large diameter seamless steel gas cylinders was studied. It is an important manufacturing process to ensure the mechanical properties of the seamless cylinder to meet the standard requirements. The influence of quenching and tempering temperature on the microstructure and properties of 34CrMo4 after complete quenching and high temperature tempering was investigated by using metallographic observation, scanning electron microscopy (SEM) and energy dispersive spectrum (EDS). The results show that the sample heat treated at an optimum process quenching at 930℃, for 60min, tempering at 560℃ for 120min can obtain the best comprehensive properties. It's beneficial to the application and safety of 34CrMo4 large diameter seamless steel gas cylinders.