ABSTRACT

 

An effective protocol was developed for in vitro propagation of ‘Virali’ via somatic embryogenesis in cell suspension culture. Embryogenic callus was obtained on Murashige and Skoog (MS) medium by a relatively moderate concentration of 2,4-dichlorophenoxyacetic acid (2,4-D) (4 mg/L), high concentrations of proline and glutamine (both 300 mg/L) and coconut water to develop induction of calli from explant. A suspension culture of the callus-derived embryogenic cells in Murashige and Skoog (MS) basal medium (pH 5.3) with 2 mg/L 2,4-D, 1 mg/L indole-3-acetic acid (IAA), 1 mg/L NAA, 45 g/L sucrose, and 20 g/L maltose resulted in synchronously proliferating cells with the capacity to be induced into somatic embryos on an 8g/L agarose,  20 g/L maltose, 10 g/L dextrose, 40 g/L sucrose, 1 mg/L naphthaleneacetic acid (NAA), and 0.2 mg/L. On an 8 g/L agarose-solidified MS basal medium (pH 5.7) with 1 mg/L NAA, 100 ml/L coconut water, 30 g/L sucrose, 30 g/L maltose, and 100 mg/L glutamine, the young somatic embryos differentiated into regenerable mature somatic embryos. In a 2.5 g/L gelrite solidified MS-based medium with 2.5 mg/L 6-benzylaminopurine (BAP), 1 mg/L gibberellic acid (GA3), 100 mg/L L-glutamine, 30 g/L sucrose, and 2.5 mg thidiazuron, the mature somatic embryos regenerated into plantlets (TDZ). For the formation of plantlets with a height of 10 to 12 cm that could endure the hardening process, it took a total of 6 months from the explant stage. The plantlets’ regular morphology suggests that there was no somaclonal variation in their development. The current regeneration approach for “Virali” has a significant deal of potential for micro propagating endangered germplasm and improving it through transgenic technology.

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Keywords   Somatic embryogenesis, Hop Bush, Virali, Dodonaea viscosa, Plant growth regulator.