Establishment of regeneration and stable transformation protocol for Eclipta prostrata (L.) L: a significant herb of Indian traditional medicinal system

The study we presented here demonstrates an efficient regeneration and transformation protocol for Eclipta prostrata (L.) L (Syn.: Eclipta alba (L.) Hassk). The establishment of regeneration protocol starts with collection of germplasm from the wild. Seeds collected from morphologically healthy plants were used as the source for in-vitro seedlings production. The optimized Murashige and Skoog (MS) medium with 4.44 μM Benzyl aminopurine (BAP) and 3% sucrose demonstrated the successful approach for generating the number of shoots (36–44 shoots/node) during the regeneration process. The altered shoots were efficiently rooted using hormone-free half MS media (HMS 20.7 g/L). The specific Agrobacterium tumefaciens strain GV3101, carrying pBI121-CFP gene (modified cyan fluorescent protein) in binary vector with kanamycin resistance, was used for plant transformation. The transformation process involved inoculating nodal explants with A. tumefaciens cells at an optical density of 0.5 and allowing an infection period of 45 min and co-cultivation for 48 h. Kanamycin, at a concentration of 35 mg/L and 45 min of infection time, was employed as a selectable condition to screen and identify successfully transformed plants. PCR analysis (T₁ and T₂ transgenic lines) and fluorescent microscopy (T₂ transgenic lines) were utilized to confirm the transgenic nature of the stable regenerated plants, ensuring successful integration of foreign genetic material. We have established a simple and stable transformation system for E. prostrata using a regeneration protocol that has promising implications. Using this protocol researchers may enhance the production of specific bioactive compounds, thereby increasing the medicinal and pharmaceutical value of E. prostrata.