The fatigue problem of orthotropic steel bridge decks of urban rail transit steel bridges has gradually become one of the hot research topics. And it is also a key problem that restricts the further development of rail transit steel bridges. In this paper, the orthotropic steel bridge deck structure of a long-span urban rail transit cable-stayed bridge is studied. Based on the segmental finite element model and full-scale model, the fatigue details of the joint weld between an orthotropic steel bridge deck and U-rib were studied theoretically and experimentally. The theoretical model of the segment is analyzed to obtain the hot spot stress characteristics. On this basis, the full-scale model fatigue test and the fatigue performance evaluation are completed based on the S-N curve. The results show that the fatigue performance of the bridge deck and U-rib joints of the orthotropic steel bridge deck structure model meets the design requirements and has a certain safety reserve. The joint fatigue details of the bridge deck, the U-rib joint weld, and the diaphragm plate are the sensitive areas that are most likely to occur fatigue failure first and need to be paid attention to in the later bridge maintenance and inspection.