Stress rupture experiments were performed on GH4151 nickel-based superalloy specimens. The microstructure analysis, fracture morphology analysis, and dislocation structure analysis were carried out. The results indicate that the life of the alloy decreased significantly with increasing temperature and stress. Furthermore, the microstructure evolution during stress rupture deformation was dislocation formed in γ channel, dislocation network, dislocation shearing into γ’ phases and stacking faults forming, extended stacking fault, and rafting γ’ phases. In high temperature, grain boundaries are easier to slide, dislocations are easy to accumulate during deformation, and the dislocations cut the γ’ phase and thus the strengthening effect is weakened, which makes the crack initiation and propagation earlier and the life of the alloy shorter. In addition, with the increasing stress, the enhancement of dislocation density and the rafting of γ’ phases becomes more obvious, which decrease strength of alloy and stress rupture life.