Genetic transformation of Tylophora indica with Agrobacterium rhizogenes A4: growth and tylophorine productivity in different transformed root clones

• Published in: Plant cell reports Vol. 24; no. 1; pp. 25 - 35
• Main Authors: Chaudhuri, K.N, Ghosh, B, Tepfer, D, Jha, S
• Format: Journal Article
• Language: English
• Published: Berlin Springer 01-04-2005
• Subjects: Agrobacterium rhizogenes; alkaloids; Alkaloids - biosynthesis; Biological and medical sciences; biosynthesis; Biotechnology; callus culture; Eukaryotic cell cultures; Fundamental and applied biological sciences. Psychology; Genetic Engineering - methods; genetic transformation; genetic vectors; indolizidine alkaloids; Indolizines; medicinal plants; Methods. Procedures. Technologies; micropropagation; Miscellaneous; molecular seqeunce data; nucleotide sequences; Phenanthrenes; Plant cells and fungal cells; plant growth; plant regeneration; Plant Roots - genetics; Plant Roots - growth & development; Plant Roots - metabolism; Plants, Genetically Modified; Rhizobium; Rhizobium rhizogenes; Time Factors; Tissue Culture Techniques; Transformation, Genetic; Tylophora; Tylophora - genetics; Tylophora - growth & development; Tylophora - metabolism; Tylophora indica; tylophorine; Medicinal plants; Vegetals; Tylophora indica; Genetic transformation; Root; Growth; Tylophorine; Bacteria; Rhizobiaceae; Agrobacterium rhizogenes; Medicinal plant
• ISSN: 0721-7714; 1432-203X
• DOI: 10.1007/s00299-004-0904-x

We have developed an efficient transformation system for Tylophora indica, an important medicinal plant in India, using Agrobacterium rhizogenes strains LBA9402 and A4 to infect excised leaf and stem explants and intact shoots at different sites. The induction of callus and transformed roots was dependent on the bacterial strain, explant type and inoculation site used. Transformed roots were induced only in explants infected with A. rhizogenes strain A4, while an optimal transformation frequency of up to 60% was obtained with intact shoots inoculated at the nodes. The presence of the left-hand transferred DNA (T(L)-DNA) in the genome of T. indica roots induced by A. rhizogenes was confirmed by PCR amplification of the rooting locus genes of A. rhizogenes. Root growth and the production of tylophorine, the major alkaloid of the plant, varied substantially among the nine root clones studied. Both parameters increased over time in liquid cultures, with maximum biomass and tylophorine accumulation occurring within 4-6 weeks of growth in fresh medium. Interestingly, in liquid culture, the culture medium also accumulated tylophorine up to concentrations of 9.78+/-0.21 mg l(-1).