Effects of plant growth regulators on efficient plant regeneration efficiency and genetic stability analysis from two Ocimum tenuiflorum L. morphotypes

Abstract

An effective, rapid and improved protocol was established for in vitro plant regeneration from nodal explants of two Ocimum tenuiflorum L. morphotypes, an medicinally important species of immense industrial value. The treatment medium used was Murashige and Skoog (MS) medium supplemented with different concentrations and combination of cytokinins (BA/KN/2-iP) and auxins (NAA/IBA/IAA). The adventitious shoot buds formation and plant regeneration capacity were significantly different (p < 0.05) between two morphotypes. The highest number of shoots (5.01 ± 0.11) in O. tenuiflorum purple leaves was obtained in the medium containing 0.5 mg l−1 BA, while in O. tenuiflorum green leaves the highest number of shoots was 7.03 ± 0.10 on a medium containing 1.0 mg l−1 BA. Regenerated shoots were rooted in vitro on low-salt MS derived media. Media were supplemented with different auxins that affected roots formation of particular morphotypes. The acclimatization of micropropagated plantlets of two morphotypes showed the best results with the sand:soil:compost (2:2:1). Genetic stability of regenerated plants as well as mother plants of two morphotypes of O. tenuiflorum L. was assessed using random amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR) markers. Both RAPD and ISSR displayed a homogenous amplification profile for all the micropropagated plants of O. tenuiflorum L. when compared to their mother plants. Gas chromatography results showed that the essential oils extracted from in vitro-derived plants and their mother stock plants have similar profiles. Rapid and high multiplication frequency, molecular genetic and essential oil content stability ensures the efficacy of the protocol developed for the production of these industrially important medicinal plants.