Establishment of Embryogenic Cell Suspension Cultures and Plantlet Regeneration in Ashwagandha (Withania Somnifera L.)

The establishment of embryogenic cell suspension culture from the embryogenic callus culture of Withania somnifera (L.) was performed by transferring mature embryo and hypocotyl embryogenic callus cultures in liquid media. The cultures obtained were swamped with clumps of proliferating globular embryos with modest non-embryogenic tissues. The number and size of somatic embryos/clumps was recorded to compute growth of embryogenic tissues under various conditions. Initiation and proliferation of embryogenic suspension culture was influenced by genotypes and various exogenous plant growth regulators fortified to the culture medium at variable magnitude. For the establishment of suspension cultures, MS medium fortified with 1.0 mgl⁻¹ 2,4-D with 0.5 mgl⁻¹ BAP was found to be the most effective. For subsequent subculturing, the reduced level of 2,4-D (0.5 mgl⁻¹) in combination with 0.5 mgl⁻¹ BAP promoted somatic embryogenesis at a faster rate. Frequent and efficient plantlet regeneration occurred on MS medium with 1.0 mgl⁻¹ BAP and 0.5 mgl⁻¹ each of NAA and kinetin. For culture efficiency genotype JA-20 responded well as compared to MWS-100. A combination of 65% relative humidity and 28^oC temperature regime exhibited higher survival of regenerated plantlets (∼90%) followed by 60% RH and 30^oC (∼85%). Later approximately 80% plants survived field transplantation.