Cellular origin and ultrastructural changes of regenerating shoots from tobacco (Nicotiana tabacum) internodes cultured in vitro

Ultrastructural changes were studied during shoot formation from tobacco internode slices cultured on Murashige and Skoog's (MS) basal medium plus 0.54 μ M naphthaleneacetic acid, 0.44 μ M 6-benzyladenine, 3% (w/v) sucrose and 0.8% (w/v) agar. Dramatic structural changes were observed in cortical cells below the internodal epidermis, especially those immediately centripetal to the stomata, by light and scanning electron microscopy. Transmission electron microscopic investigation revealed conspicuous structural changes to plastids at each stage during shoot regeneration. To confirm the significance of the cortical cells in shoot regeneration, protoplasts were isolated from them and shoots regenerated successfully. The ability of single cortical cell protoplasts to differentiate into shoots was demonstrated.