Stable vindoline production in transformed cell cultures of Catharanthus roseus

Catharanthus roseus L. (G. Don) leaf disks were transformed by cocultivation with seven strains of Agrobacterium tumefaciens and Agrobacterium rhizogenes. Four distinct transformed cell lines were. obtained with A. tumefaciens strains A281, BO542, and A. rhizogenes strain K599, and selected on hormone-free medium. Transformation of the cell lines was confirmed by Southern blot analysis, and preliminary data on the alkaloid profiles was determined by 2D TLC and HPLC/MS. Stable vindoline production was demonstrated in the undifferentiated suspension cell lines strains, CR-BO542, CR-K599, and strain CR-A281S (shooty teratomas), by HPLC/MS analysis of the alkaloid fractions. Active vindoline biosynthesis in the suspension-cell lines was further confirmed by measuring the activity of acetyl-coenzyme-A-deacetylvindoline-O-acetyltransferase, one of the final enzymes in the biosynthetic pathway of vindoline, in cell-free extracts.