Production of genetically uniform plants from shoot tips of Aconitum heterophyllum Wall. – a critically endangered medicinal herb

Abstract

ABSTRACTWe report the successful micropropagation of a critically endangered medicinal plant Aconitum heterophyllum Wall., using low concentrations of plant growth regulators (PGRs) and molecular validation of the clonal stocks. The maximum rate of in vitro shoot multiplication was obtained on 1.0 × Murashige and Skoog (MS) medium containing 0.25 mg L−1 Kinetin (Kn) plus 0.25 mg L−1 Indole acetic acid (IAA). Up to 100% rooting was obtained 15 for shoots cultured on 1.0 × MS medium supplemented with 1.0 mg L−1 IAA. Adding 0.25 mg L−1 2,4-dichlorophenoxyacetic acid (2, 4-D) to 1.0 × MS medium resulted in 100% callus formation, while adding 0.25 mg L−1 IAA plus 0.25 mg L−1 Kn to 1.0 × MS medium containing 0.25 mg L−1 2,4-D resulted in 100% generation of embryogenic callus. Inter-simple sequence repeat (ISSR) marker analysis was carried out to check for possible somaclonal variation in the plantlets obtained after three consecutive sub-cultures. Of the 15 ISSR primers used, 10 were found to be monomorphic, with 95–98% similarity, and were used for cluster analysis by the unweighted pair group using arithmetic averages (UPGMA) method. The results revealed that in vitro-regenerated plantlets did not exhibit any genetic polymorphism.