Examination of the Conditions for Protoplast Isolation from Tobacco Cells Cultured in vitro

Protoplasts were isolated from cultured tobacco cells by removing the cell wall enzymatically. We examined the time courses of treatment with some different concentrations of enzymes (cellulase and macerozyme) and with some different kinds and concentrations of sugars (sucrose, sorbitol and mannitol) which produce good conditions for protoplast isolation. The best conditions for protoplast preparation from tobacco cells cultured in vitro were: use of actively growing cells and isolation of protoplasts with 5% cellulase and 0.1% macerozyme in 0.5% sorbitol.