The main span steel box girder of Jiu-jiang highway bridge over Yangtze river is studied in this paper.Through finite element analysis on the real bridge structure,the fatigue specimen which can reflect stress state of rib-to-reck welded joints of orthotropic steel bridge deck is made.Based on the theoretical analysis and static load test;the corresponding relationship of the load and stress of research part is gotten.The fatigue failure form of the structure detail,stress-life curve and allowable stress amplitude are obtained by the fatigue test.The result can be used for the evaluation of fatigue life,health detection and maintenance and repair of the bridge,which can also be used as reference for the design of other bridges.