Development of an efficient in vitro system for micropropagation and regeneration of Justicia adhatoda, Sida acuta, Pimenta dioica, and Premna integrifolia

Abstract Medicinal plants have always been a non-exhaustible source of therapeutics throughout human history. Meanwhile, their in situ populations are under increasing anthropogenic pressure, specifically overharvesting and habitat destruction. Plant tissue culture is a powerful tool in biotechnological strategies of controlled selection, ex situ preservation and mass cloning of elite genotypes in medicine. The current study represents a study focusing on the development of an effective in vitro culture protocol of four important medicinal plant species: Justicia adhatoda , Sida acuta , Pimenta dioica , and Premna integrifolia . Induction of callus was achieved successfully by adding different concentrations of auxins and cytokinins to basal growth medium, especially 2,4-dichlorophenoxyacetic acid and Thidiazuron. Maximum callus formation was achieved in J. adhatoda (76.67%), S. acuta (88.33%), P. dioica (68.33%), and P. integrifolia (81.66%). Shoot regeneration was significantly enhanced by 6-benzylaminopurine, with responses of 62.22%, 73.33%, 65.55%, and 77.78% in the respective species. Root induction was effectively obtained with a combination of auxins, showing 72.22% in P. integrifolia , 79.99% in S. acuta , 84.44% in J. adhatoda , and 76.66% in P. dioica . The regenerated plantlets were acclimatized successfully under greenhouse conditions, confirming their survival and adaptability. The protocol showed very high efficiency in callus induction, shoot proliferation and root formation and therefore a valuable tool to be used in conservation, genetic improvement and sustained commercial exploitation of these medicinal species.