Efficient in Planta Genetic Transformation of Tomato Seeds (cv. Anand Roma) through Piercing and Agrobacterium-Mediated Incubation

Tomato (Solanum lycopersicum L.) is a globally important crop and a model plant for genetic and biotechnological studies. Conventional tissue culture-based transformation is often limited by genotype dependency, high cost and somaclonal variation. In this study, an optimized in planta Agrobacterium-mediated transformation protocol was developed for tomato seeds cv. Anand Roma using the binary vector pCAMBIA 1301. Competent Agrobacterium tumefaciens (EHA105) cells harboring the construct were prepared and confirmed by colony PCR with hptII gene-specific primers. The results showed that transformation efficiency peaked at 200 mM acetosyringone (23%), 30 min soaking in Agrobacterium culture (17%), 48 hrs dark incubation (14%), and 30 mg/L hygromycin (20%). Molecular analysis of putative transformants confirmed stable integration of the hptII gene. This protocol offers a simplified, cost-effective, and genotype-specific transformation approach, providing a reliable platform for tomato improvement programs and functional genomics studies while reducing reliance on tissue culture regeneration.