Abstract: The CRISPR/Cas-mediated gene editing technology enables the precise cleavage of target DNA sequences, overcoming the limitations of traditional genetic modification methods such as low efficiency, poor specificity, and operational complexity. It has significantly simplified the gene editing process and is now widely applied in medicine, agriculture, and basic research. Although CRISPR/Cas-based gene editing has achieved remarkable progress in various crops, its application in tobacco remains relatively underdeveloped, with challenges such as low editing efficiency and suboptimal target specificity, limiting its broader application in tobacco genetic improvement. Therefore, this review focuses on introducing the principles and methodologies of gene editing technology, tracing its early development, and exploring in depth the working mechanisms, structural classifications, and applications of the CRISPR/Cas system in genome editing. The article also briefly discusses the applications of gene editing in improving tobacco resistance to diseases, insect and experimental stresses, as well as in enhancing quality traits, agronomic performance , biofuel production, and environmental remediation. Finally, it outlines the current prospects and challenges in tobacco variety improvement. Through these discussions, the study aims to provide researchers with a comprehensive overview of gene editing technologies and to offer insights and references for tobacco breeding.