Growth and terpenoid indole alkaloid production in Catharanthus roseus hairy root clones in relation to left- and right-termini-linked Ri T-DNA gene integration

Abstract

Hairy root cultures of Catharanthus roseus var. Prabal were established by infecting the leaves with Agrobacterium rhizogenes agropine-type A4 strain. Two hundred and fifty independent root clones were evaluated for growth, morphology, number of integration of Ri T-DNA genes and alkaloid contents. On the basis of growth pat- tern, type of branching and number of lateral roots we were able to separate the hairy root clones into four categories. However based on the integration of the Ri TL-DNA and TR-DNA genes, there were only three different categories of independent hairy root clones–C1 (rolA&B+/ags+), C2 (rolA&B-/ags+) and C3 (rolA&B+/ags ). Southern hybridization analysis revealed both single and multiple copies of T-DNA integration in the root clones. The accumulation of considerable amounts of the root-specific alkaloids ajmalicine and serpentine was observed in the presence of both the TL-DNA and TR-DNA genes (C1) and the TL-DNA gene (C3) alone. Two rolA&B but ags+ clones (C2) accumulated much less or only very negligible amounts of ajmalicine. The possible role of the TL-DNA and TR-DNA genes on growth and alkaloid accumulation in these root clones is discussed