Effect of cytokinin and MS medium composition on efficient shoot proliferation of Nyctanthes arbor-tristis L. through cotyledonary node explant and evaluation of genetic fidelity and antioxidant capacity of regenerants

Abstract

Nyctanthes arbor-tristis L. is an important member of family Oleaceae. An efficient shoot proliferation response (21.53±0.58 shoots/explant) from cotyledonary node was achieved on Murashige and Skoog (MS) medium supplemented with 22.19 µM 6-benzyl amino purine (BAP). To optimize the role of ammonium and nitrate on shoot proliferation, the explants were cultured on modified MS media (MS-1 to MS-12). Among these media, MS-3 and MS-9 were found most suitable for shoot-proliferation than others. The microshoots after indole 3-butyric acid (IBA) pulse treatment (1968.11 µM) for 30 min, when sub-cultured on ½ strength MS basal medium were successfully rooted with a maximum frequency of root induction (77.66±1.45), the maximum number of roots/ shoot (10.66±0.88) and maximum root length (1.60±0.05 cm). In-vitro rooted plants were acclimatized and transplanted outside into field with 80 percent survival-rate after 90 days. The genetic-fidelity was confirmed by monomorphic nature of inter-simple sequence repeat (ISSR) marker pattern. The ethanol extract of leaves of regenerated plants contained good amount of phenol (118.00±0.82 µg/mg gallic acid equivalent) and flavonoid (17.34±0.64 µg/mg rutin equivalent) and possess strong antioxidant activity. The EC50 value for 2, 2-diphenyl-1-picrylhydrazyl (DPPH) free radical, superoxide radical (SOD) scavenging activity and lipid peroxidation (LPO) inhibition activity was 339.98±3.27, 755.80±75.50 and 689.67±2.46 µg/ml respectively. Hence, these findings suggest that regenerated plants can be used as a therapeutic and inhibitory agent against free radical-induced disease progression.