Genetic transformation in ashwagandha (Withania somnifera (L.) Dunal) for hairy root induction and enhancement of secondary metabolites

Smini Varghese, R. Keshavachandran, Bincy Baby, P.A. Nazeem

Abstract


Genetic transformation was carried out in ashwagandha (Withania somnifera (L.) Dunal) using three different Agrobacterium rhizogenes strains viz., A4, ATCC 15834 and MTCC 2364, for inducing hairy roots. The explants such as hypocotyls, cotyledonary segments, leaf segments, shoot tips and nodal segments were used for genetic transformation. A4 and ATCC 15834 strains produced successful transformation and hairy (transformed) roots were induced from leaf segments and shoot tips. A4 strain produced transformation by direct inoculation of bacteria from single cell colonies as well as in the suspension form, but ATCC 15834 produced transformation only in the suspension form. Among the liquid media tested, half MS was found to be superior in promoting hairy root growth. The transformation was confirmed by PCR and dot blot analysis. A Thin Layer Chromatographic method was employed for withanolide estimation. The spot corresponding to withaferin A was observed under UV at 254 nm. Field root possessed more withaferin A followed by hairy roots and in vitro roots contained the least. Enhancement of secondary metabolite production was attempted through addition of osmoregulator, precursor feeding and elicitation. Withaferin A content in the hairy root biomass and the culture medium were estimated. The biotic elicitor Aspergillus homogenate (250 and 500 μl /125 ml) had a positive influence in the enhancement of secondary metabolites

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Keywords


Genetic transformation, Ashwagandha, Hairy root

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A KAU publication [CODEN: JTAGEI; ISSN 0971-636X; eISSN 0973-5399]