Recent advances in CRISPR-Cas9-based genome insertion technologies have significantly enhanced the precision and efficiency of targeted DNA insertion in both basic research and therapeutic applications. This review summarizes the latest developments in programmable genome insertion techniques, including their mechanisms, advantages, limitations, and potential applications. The key approaches discussed include homology-directed repair (HDR), non-homologous end-joining (NHEJ), microhomology-mediated end-joining (MMEJ), and single-strand annealing (SSA). HDR is the most commonly used method for targeted genome insertion, relying on homologous recombination to achieve precise insertions. NHEJ, while more efficient, often results in insertions or deletions (indels) at the DSB junctions. MMEJ and SSA offer alternative pathways for targeted insertion with varying efficiencies. Recent advancements have introduced new tools such as prime editing, integrase-coupled programmable nucleases, and CRISPR-associated transposons (CAST), which enable precise and efficient genome editing without generating DSBs. Prime editing allows for precise insertions and substitutions, while CAST systems, including subtype I-F and V-K, facilitate site-specific DNA integration without relying on DNA repair pathways. These technologies have been applied in various contexts, including the correction of genetic disorders, the development of cell models, and the improvement of gene therapy approaches. Despite these advancements, challenges remain, such as the need for higher efficiency, reduced off-target effects, and the ability to insert larger DNA fragments. Future research aims to enhance the precision and safety of these technologies, enabling their broader application in both research and clinical settings. The integration of these tools into therapeutic strategies, such as CAR-T cell engineering, highlights their potential to improve the safety and efficacy of gene-based therapies. Overall, the continuous development of programmable genome insertion technologies is crucial for advancing genetic research and therapeutic applications.Recent advances in CRISPR-Cas9-based genome insertion technologies have significantly enhanced the precision and efficiency of targeted DNA insertion in both basic research and therapeutic applications. This review summarizes the latest developments in programmable genome insertion techniques, including their mechanisms, advantages, limitations, and potential applications. The key approaches discussed include homology-directed repair (HDR), non-homologous end-joining (NHEJ), microhomology-mediated end-joining (MMEJ), and single-strand annealing (SSA). HDR is the most commonly used method for targeted genome insertion, relying on homologous recombination to achieve precise insertions. NHEJ, while more efficient, often results in insertions or deletions (indels) at the DSB junctions. MMEJ and SSA offer alternative pathways for targeted insertion with varying efficiencies. Recent advancements have introduced new tools such as prime editing, integrase-coupled programmable nucleases, and CRISPR-associated transposons (CAST), which enable precise and efficient genome editing without generating DSBs. Prime editing allows for precise insertions and substitutions, while CAST systems, including subtype I-F and V-K, facilitate site-specific DNA integration without relying on DNA repair pathways. These technologies have been applied in various contexts, including the correction of genetic disorders, the development of cell models, and the improvement of gene therapy approaches. Despite these advancements, challenges remain, such as the need for higher efficiency, reduced off-target effects, and the ability to insert larger DNA fragments. Future research aims to enhance the precision and safety of these technologies, enabling their broader application in both research and clinical settings. The integration of these tools into therapeutic strategies, such as CAR-T cell engineering, highlights their potential to improve the safety and efficacy of gene-based therapies. Overall, the continuous development of programmable genome insertion technologies is crucial for advancing genetic research and therapeutic applications.