Regeneration efficiency was enhanced by adding auxin and cytokinin at different concentrations to the media. It was found that the two highest frequencies-90 and 85%-of callus induction occured when the buds were cultured in an MS medium supplemented with 2 mgL-1 α-naphthalene acetic acid (NAA) plus 1 mgL-1 N 6-benzylaminopurine (BAP) (CIM2) and in the same medium supplemented with 2 mgL-1 NAA plus 1 mgL-1 kinetin (CIM5), respectively. When cultured further in a plant regeneration medium supplemented with 1 mgL-1 NAA and 2 mgL-1 BAP (PRM1), these calli regenerated with high efficiencies, 80% for calli grown in CIM2 and 85% for calli grown in CIM5. The low BAP concentration in CIM1 (2 mgL-1 NAA and 0.5 mgL-1 BAP) resulted in a low regeneration efficiency and fewer shoots per callus even though the PRM3 regeneration medium had a high concentration of cytokinins (2 mgL-1 BAP and 2 mgL-1 kinetin). However, the calli cultured in CIM2 without kinetin but with a suitable level of BAP still regenerated with high efficiencies in PRM2 and PRM3 that contained 1 and 2 mgL-1 kinetin, respectively. Similarly, when calli cultured in CIM5 without BAP but with kinetin were regenerated in PRM1 with BAP alone, they also regenerated with a high regeneration efficiency. These results showed that culturing a specific tissue in a medium with suitable concentrations of NAA and BAP or kinetin resulted in a large number of multiple shoots, and BAP and kinetin worked synergistically to produce a high plant regeneration efficiency.

Zinc is an essential micronutrient element for plant. Nowadays, Zinc oxide nanoparticles (ZnO NPs) are widely used and have been shown adverse effects to plant physiology. The optimal concentration of ZnO NPs added in plant regeneration medium (PRM) is studied in this experiment. Six-weeks old calli induced from the aseptic axillary buds of vetiver grass were cultured in PRM with various concentrations of ZnO NPs (5, 10, 20, 40 mgL-1). PRM containing 5, 10 mgL-1 ZnO NPs showed the highest percentage of plant regeneration frequency (95%) but did not have a statistic difference with other treatments. However, the averages of shoot length regenerated under both concentrations were 1.32-1.33 cm. These averages of shoot length had statistically different from averages of shoot length of the others. Moreover, calli regenerated under PRM containing 5 mgL-1 ZnO NPs condition showed the highest average number of shoots per callus (9.75 shoots) with a statistic difference. ZnO NPs at 20-40 mgL-1 in the PRM did not enhance the percentage of plant regeneration, but retarded calli development and shoot elongation. The optimal concentration of ZnO NPs in PRM has positive effects on regeneration and development of vetiver grass under tissue culture condition.

The optimum ratios of auxin and cytokinin are necessary for callus induction and plant regeneration. This ratio is a key function involving in the promoting cell division and proliferation in tissue culture. The axillary buds of in vitro plantlets fromVetiveria nemoralisA. Camuscv. Roiet were used as explants for the callus induction experiment. These explants were cultured on Murashige & Skoog (MS) medium [1] supplemented with various combinations of auxins and cytokinins. Under this experimental study, the highest frequency of callus induction was found on MS medium supplemented with 2 mgL-1α-naphthalene acetic acid (NAA) and 1 mgL-12-furanylmethyl-1H-purine-6-amine (kinetin) (62.5%). On the other hand the combination of 2, 4-dichlorophenoxyacetic acid (2, 4-D) and 6-benzylaminopurine (BAP) was toxicity to this explants. All culturing explants were dead and no calli appearance. The calli derived from each medium were transferred into the same regeneration medium (MS with 1 mgL-1NAA and 2 mgL-1BAP). After culturing on regeneration medium, calli induced from the highest callus induction medium have shown high frequencies of regeneration and also shoot number per callus (58.33% and 7.1 shoots).