AGROBACTERIUM RHIZOGENES-MEDIATED TRANSFORMATION AND PLANT-REGENERATION OF 4 GENTIANA SPECIES

Shoots of micropropagated Gentiana acaulis, G. cruciata, G. lutea, and G. purpurea were inoculated with suspensions of Agrobacterium rhizoge nes cells, strains ATCC 15834 or A4M70GUS. Adventitious roots appeared at the sites of inoculation in all 4 species. Root tips were excised and cultured on growth regulator-free media for 2-6 years. They exhibi ted very high branching and plagiotropism. Spontaneous bud initiation occurred in roots of G. cruciata. Roots of G. lutea, G. acaulis and G. purpurea were cultured on media with high kinetin concentration, whic h induced the formation of friable callus tissues. Only in G. purpurea were these calluses organogenic. Regenerated shoots of G. cruciata an d G. purpurea gave rise to plants, that displayed the typical phenotyp es of A. rhizogenes-transformed plants: short internodes and rolled le aves. In the roots of G. acaulis and G. cruciata, transformed with A. rhizogenes A4M70GUS, a positive reaction with X-gluc indicated the act ivity of beta-glucuronidase. The DNA extracted from hairy roots and fr om the roots of transgenic plants hybridized with the appropriate geno mic probes in Southern blotting. This is taken as evidence of the stab le genetic transformation in the 4 Gentiana species.