Long-term survival of Vicia guard cell protoplasts in cell culture

Abstract

Guard cell protoplasts from Vicia faba L. survived for 40 days in culture under aseptic conditions in a minimal medium containing 0.45 M mannitol and 1 mM CaCl 2. Forty days after plating, protoplasts kept in 8 h of 100 μmol m −2 s −1 red light/16 h darkness per day had a 50% survival rate, in contrast with a 5% survival rate of cells kept in darkness. In 12-day long experiments, 80% of the guard cell protoplasts survived both under red light (8 h of 50 μmol m −2 s −1) and in darkness. In contrast, 45% of mesophyll protoplasts survived in the light and 25% in darkness. Cultured guard cell protoplasts exhibited normal, blue light-stimulated swelling for at least 1 week. Addition of salts and sugars did not increase protoplast survival. Some morphogenetic responses, including wall formation, budding and incipient cytokinesis, but not cell division, were observed under different culture conditions. Cell culture of Vicia guard cell protoplasts can provide a long-term source of cells for physiological experiments and an experimental system for the study of cell differentiation.