Abstract: Single-atom catalysts (SACs), as the rising stars in the field of catalytic science, are leading catalytic technology into an unprecedented new era. However, the synthesis of high-performance SACs with well-defined active sites and high loadings under precise control has become a hotly debated topic in scientific research. Metal-organic frameworks (MOFs), with their exceptional properties such as ultrahigh specific surface areas, precisely controllable structural designs, and highly flexible functional customization capabilities, are regarded as one of the ideal matrices for supporting and stabilizing SACs. This review provides an insightful overview of the diverse preparation strategies for MOFs-derived SACs. It comprehensively analyzes the unique advantages and challenges of each method in achieving efficient synthesis of SACs, emphasizing the crucial role of optimized processes in unlocking the anticipated performance of SACs. Furthermore, this review delves into a series of advanced characterization techniques, including aberration-corrected scanning transmission electron microscopy (AC-STEM), electron energy loss spectroscopy (EELS), X-ray absorption spectroscopy (XAS), and infrared absorption spectroscopy (IRAS), offering valuable insights into the atomic-scale fine structures and properties of SACs, significantly advancing the understanding of SAC mechanisms. Moreover, this review focuses on exploring the potential applications of MOFs-derived SACs in electrocatalysis frontier fields. This comprehensive examination lays a solid theoretical foundation and provides a directional guidance for the rational design and controllable synthesis of high-performance MOFs-derived SACs.