Genotype-flexible plant genetic transformation: advances and prospects

Genetic transformation of elite crop varieties remains limited by genotype-specific recalcitrance and dependence on tissue culture. This review provides a comprehensive analysis of current transformation platforms and emerging strategies to overcome these bottlenecks. We examine traditional <i>Agrobacterium</i>-mediated and biolistic methods, then critically assess tissue culture-free approaches including floral-based delivery, cut-dip-budding, <i>in planta</i> injection, and viral vector-mediated transformation systems. A major focus is the deployment of developmental regulators-<i>WUS/BBM</i>, <i>GRF-GIF</i> chimeras, <i>DOF</i> transcription factors, <i>LAX1</i>, <i>PLT</i>, and <i>WIND1</i>-that enhance regeneration efficiency across genotypes. We detail their molecular mechanisms, from chromatin remodeling and auxin gradient establishment to wound-responsive cell reprogramming. Importantly, we address the pleiotropic developmental costs of DR misexpression and review precision control technologies, including promoter optimization and auto-excision systems, that enable transient DRs activity during regeneration while ensuring normal plant development. We propose a roadmap for integrating these advances to achieve genotype-flexible, high-throughput transformation applicable to molecular design breeding.(1).