Metabolic‐Engineering Approach to Enhance Vanillin and Phenolic Compounds in Ocimum Sanctum (CIM‐Angana) via VpVAN Overexpression

AbstractTransgenic Ocimum sanctum plants were engineered to produce vanillin by overexpressing the VpVAN gene using Agrobacterium‐mediated transformation. Positive transformants developed shoots within 4–5 weeks and were transferred to a root induction medium and four independent transformants with no observed adverse effects were kept for anlysis. Quantitative RT‐PCR indicated significantly higher VpVAN expression in transgenic lines AG_3 and AG_1, impacting the phenylpropanoid pathway and phenolic compound accumulation. Molecular docking studies indicated ferulic acid's higher binding affinity to vanillin synthase than eugenol. LC–MS/MS analysis revealed a marked increase in vanillin production in transgenic lines compared to wild type, with AG_3 exhibiting the highest vanillin content (1.98 ± 0.0047 mg/g extract) and AG_1 following (1.49 ± 0.0047 mg/g extract). AG_3 also showed elevated levels of benzoic acid, 4‐hydroxy benzyl alcohol, and ferulic acid. This study highlights the potential of metabolic engineering in O. sanctum for enhanced vanillin production, suggesting pathways for large‐scale production of natural vanillin and other valuable compounds in transgenic plants.