Rhizobium rhizogenes Mediated Hairy Root Transformation and Analysis of Secondary Metabolites in Phlogacanthus thyrsiflorus Nees Hairy Roots using GC-MS

An efficient transformation system was developed for Phlogacanthus thyrsiflorus, a medicinal plant of northeast India, using Rhizobium rhizogenes strains A4 for infecting shoot tip, nodes and leaf explants derived from aseptically raised plants. The transformation frequency was found dependent on the explants as well as the co-culture periods. Highest transformation frequency of 95.69% was obtained from shoot tip explants co-cultured for 4 hours. An average of 6.37 hairy roots per explant were obtained from shoot tip explants within three weeks of infection. Whereas, transformation frequency along with number of transformed roots were fewer in node and leaf explants. The presence of T-L DNA of R. rhizogenes strains A4 in the hairy root genome of P. thyrsiflorus was confirmed through PCR based cloning and sequencing in which the sequencing results of the PCR amplicon confirmed the presence of rol b and rol c genes in the host genome. Sequence results for rol genes were found to be 97 percent and 99 percent similar with rol b and rol c gene present in NCBI database. GC-MS analysis of methanolic extract of the hairy roots reveals the presence of a few medicinally important secondary metabolites.