Abstract: High-cycle fatigue tests on SUS301L stainless steel Laser-MAG hybrid welded butt joint were conducted with considering the effect of stress ratio. The fatigue life of welded joint was evaluated by using the hot spot stress method recommended by the IIW code. The experimental analysis and research results show that the stress ratio has a significant effect on the fatigue life of the welded joint specimens, and the fatigue life increases with the increase of the stress ratio. The design curves based on the hot spot stress method and stress ratio modification methods can not be applicable to fatigue life evaluation of the SUS301L stainless steel welded joint. The effect of stress ratio on the fatigue life of welded joint can be reasonably reflected by using the Walker’s equivalent hot spot stress.