A Revised Medium for Growth of Pea Mesophyll Protoplasts

The nutrient requirements of mesophyll protoplasts from Pisum sativum L. cv. Timo have been investigated and a synthetic and completely defined medium has been designed. A high calcium concentration (12 mM) stimulated both protoplast survival and cell division. The content of iron and zinc was also critical. Additions of nicotinic acid, pyridoxine and thiamine were necessary. The protoplast growth was enhanced when some amino acids were included in the medium. An absolute requirement for auxin and cytokinin was shown. In the revised medium about 90% of the isolated protoplasts survived and formed a cell wall. The first divisions were observed after 5 days and after 1 week 10–20% of the cells had divided at least once.