title: Establishment of Withania somnifera hairy root cultures for the production of withanolide A.
abstract: Withania sominifera (Indian ginseng) was transformed by Agrobacterium rhizogenes. Explants from seedling roots, stems, hypocotyls, cotyledonary nodal segments, cotyledons and young leaves were inoculated with A. rhizogenes strain R1601. Hairy (transformed) roots were induced from cotyledons and leaf explants. The transgenic status of hairy roots was confirmed by polymerase chain reaction using nptII and rolB specific primers and, subsequently, by Southern analysis for the presence of nptII and rolB genes in the genomes of transformed roots. Four clones of hairy roots were established; these differed in their morphology. The doubling time of faster growing cultures was 8-14 d with a fivefold increase in biomass after 28 d compared with cultured, non-transformed seedling roots. MS-based liquid medium was superior for the growth of transformed roots compared with other culture media evaluated (SH, LS and N6), with MS-based medium supplemented with 40 g/L sucrose being optimal for biomass production. Cultured hairy roots synthesized withanolide A, a steroidal lactone of medicinal and therapeutic value. The concentration of withanolide A in transformed roots (157.4 microg/g dry weight) was 2.7-fold more than in non-transformed cultured roots (57.9 microg/g dry weight).
authors: Plant Sciences Division, School of Biosciences, University of Nottingham, Sutton Bonington Campus, Loughborough, LE12 5RD, UK. nmurthy60@yahoo.co.in
authors: Hosakatte N
authors: HN
authors: Murthy
authors: Camelia
authors: C
authors: Dijkstra
authors: Paul
authors: P
authors: Anthony
authors: Daniel A
authors: DA
authors: White
authors: Mike R
authors: MR
authors: Davey
authors: J Brian
authors: JB
authors: Power
authors: Eun J
authors: EJ
authors: Hahn
authors: Kee Y
authors: KY
authors: Paek
mesh_terms: D015139:Blotting, Southern; D018929:Cell Culture Techniques / Q000379:methods*; D002851:Chromatography, High Pressure Liquid; D003470:Culture Media; D004269:DNA, Bacterial / Q000032:analysis; D004875:Ergosterol / Q000031:analogs & derivatives* / Q000032:analysis / Q000096:biosynthesis; D005798:Genes, Bacterial; D018515:Plant Leaves / Q000382:microbiology; D018517:Plant Roots / Q000166:cytology* / Q000254:growth & development / Q000378:metabolism*; D030821:Plants, Genetically Modified; D016133:Polymerase Chain Reaction; D012015:Reference Standards; D012231:Rhizobium / Q000235:genetics; D014170:Transformation, Genetic; D032341:Withania / Q000166:cytology* / Q000235:genetics / Q000378:metabolism*; D054358:Withanolides
chemical_list: C433551:3-rhamnopyranosyl(1-4)-glucopyranosyl-12-diacetoxy-20-hydroxywitha-5,24-dienolide; D003470:Culture Media; D004269:DNA, Bacterial; D054358:Withanolides; D004875:Ergosterol