The orthotropic steel bridge deck weld structure would easily cause fatigue cracking under the repeated action of vehicle load. This paper took the steel box girder in a bridge as a research object, researched the mechanical properties of the steel plate and the microstructure of the welded joint, then designed the fatigue specimens of the deck plate and did the fatigue test. The Δσ-N curves and stress amplitudes of the weld details of the deck plate with U-rib and diaphragm under different probabilities of survival were obtained. After extended the Δσ-N curves to the long life range, the fatigue damage calculation equation of the detail was proposed, and the cut-off limit under the 50% and 97.7% probability of survival were 81.50 MPa and 53.11 MPa, respectively. Based on the actual vehicle load spectrum and simplified finite element model of the steel box girder section, the stress amplitude of the details of the weld joint was calculated. The calculation result shows that the maximum stress amplitude of the concerned point was 38.29 MPa, less than the cut-off limit. It means that the fatigue strength of the details of the weld joint meet the requirement of the fatigue design.