Manipulation of root biomass and biosynthetic potential of Glycyrrhiza glabra L. plants by Agrobacterium rhizogenes mediated transformation

Abstract

Agrobacterium  rhizogenes  is  gram­negative  soil  bacteria  and  the  causal  agent  for  the development  of the  hairy  root  disease in  plants.  Hairy  roots  cultures  have  been  proven  as  being  a natural  source for  secondary metabolites that  are  normally  biosynthesized in  roots  of  differentiated plants. Here we established an experimental condition for obtaining hairy roots culture of medicinal plant  Glycyrrhiza  glabra  as  effect  of  integration  and  expression  of rolB  gene  from  Ri  plasmid  of A.rhizogenes. Two Agrobacterium strains, LBR56 and LBA9402 were tested for hairy roots induction but only LBR56 strain show an increase in transformation efficiency relative to wild­type. This high productive  root  biomass  was  induced  from  in  vitro  old  roots  and  internodes  segments  by  direct infection with A. rhizogenes LBR56 strain. Several independent hairy roots clones were obtained and maintained in vitro and further evaluated for growth, morphology, integration and expression of Ri T­ DNA  genes.  The  PCR  analysis  with  specific  primers  for  the  rol  B  gene  and  nptII  marker  gene confirmed the integration  of  T­DNA fragment  of  Ri  plasmid from  A.rhizogenes into the  genome  of G.glabra transgencic plants. RT­PCR analysis confirmed the expression of hairy roots inducible gene. The  glycyrrhizic  acid  (GA)  yield  has  been  monitored  in  the  hairy  roots  biomass  by  UV  spectral analysis.