Effect of aminoglycoside antibiotics on in-vitro morphogenesis from cultured cells of chrysanthemum and tobacco

Successful genetic transformation of plants requires non-chimeric selection of transformed tissues and their subsequent regeneration. With rare exceptions, most transformation protocols still rely heavily on antibiotics for selecting transgenic cells that contain an antibiotic-degrading selectable marker gene. Here, the morphogenic capacity of in-vitro explants of chrysanthemum and tobacco stems and leaves (control and transgenic) changed with the addition of aminoglycoside antibiotics (AAs). In a test of 6 AAs, phytotoxicity occurred at concentrations of 10 to 25 and 50 to 100 mug mL(-1) in chrysanthemum and tobacco explants,. respectively. Light conditions as well as explant source and size also had significant effects. The use of transverse thin cell layers (tTCLs), in conjunction with high initial AA selection levels, supported the greatest regeneration of transgenic material (adventitious shoots or callus) and the lowest number of escapes. Flow-cytometric analyses revealed no endoduplication in chrysanthemum, even at high AA levels. However, this phenomenon was observed in tobacco calli (8C or more), even at low AA concentrations (i.e., 5 to 10 mug mL(-1)).