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+ Recent progress in personalized image generation using diffusion models has been significant. However, development in the area of open-domain and non-fine-tuning personalized image generation is proceeding rather slowly. In this paper, we propose Subject-Diffusion, a novel open-domain personalized image generation model that, in addition to not requiring test-time fine-tuning, also only requires a single reference image to support personalized generation of single- or multi-subject in any domain. Firstly, we construct an automatic data labeling tool and use the LAION-Aesthetics dataset to construct a large-scale dataset consisting of 76M images and their corresponding subject detection bounding boxes, segmentation masks and text descriptions. Secondly, we design a new unified framework that combines text and image semantics by incorporating coarse location and fine-grained reference image control to maximize subject fidelity and generalization. Furthermore, we also adopt an attention control mechanism to support multi-subject generation. Extensive qualitative and quantitative results demonstrate that our method outperforms other state-of-the-art frameworks in single, multiple, and human customized image generation. Please refer to our project's website.