To investigate the temperature distribution of box girder, an experiment is conducted to reveal the temperature distribution of a section of Fenghe Bridge, which is a single box three-room concrete bridge located in the Xixian New District of Xi’an City. Experimental results show that the concrete temperature of outside surface was greatly affected by the ambient temperature that fluctuates rapidly every day, but the temperature of inside surface was relatively constant except the top plate. From the viewpoint of vertical temperature distribution, the one-dimensional heat transmission theory is suitable for the upper and bottom flange, but not for the web. Meanwhile, the temperature difference between outside and inside bottom flange is obvious. In addition, the maximum gradient of positive temperature mainly appeared in summer, while the maximum gradient of negative temperature mainly appeared in winter. According to the experimental data, the temperature distribution pattern and temperature base of box girder were discussed, and a new temperature gradient function has been proposed by combing with exponential and linear functions.