Efficient plantlet regeneration via somatic embryogenesis and shoot organogenesis in Dichanthium annulatum (Forssk.): a halophyte C4 apomictic forage crop

Dichanthium annulatum is a popular halophyte C-4 perennial forage crop of semi-arid tropics that reproduces apomictically. In vitro plant regeneration in apomictic species helps in producing new germplasm that can supplement existing breeding program. Field-collected seeds and in vitro shoot apices were tested for response to 2,4-dichlorophenoxyacetic acid (2,4-D) and 6-benzylaminopurine (BAP) for initiation of somatic embryogenesis (SE) and direct shoot organogenesis (DSO). Explants produced callus on Murashige and Skoog (MS) medium with varying level of 2,4-D and 0.5 mgL(-1) BAP. After 4th sub-culture, yellowish brown turns into white nodular callus tissue with a friable appearance, which exhibited the highest percentage of embryogenic callus induction on MS medium with 3.0 mgL(-1) 2,4-D and 0.5 mgL(-1) BAP, was transferred to MS with BAP or kinetin and 0.125 mgL(-1) 2,4-D to promote somatic embryo germination into plantlet. The developmental stages of SE were identified using scanning electron microscopy and histology. Multiple shoots were observed with the combinations of BAP or kinetin with 2,4-D. The highest number of shoots (27) occurred on MS medium comprising 4.0 mgL(-1) BAP and 0.125 mgL(-1) 2,4-D. In vitro plantlet was rooted on MS medium with 0.4% charcoal. The rooted 80% plants could be acclimatized and established in soil, as they showed healthy growth and fertility.