Sustained division of protoplast-derived cells from primary leaves of Pinus pinaster, factors affecting growth and change in nuclear DNA content
- 1 June 1986
- journal article
- research article
- Published by Oxford University Press (OUP) in Tree Physiology
- Vol. 1 (1) , 21-30
- https://doi.org/10.1093/treephys/1.1.21
Abstract
Leaf protoplasts were isolated from apical and in vitro-induced axillary buds of Pinus pinaster Ait. seedlings. First divisions were seen after 8–10 days of culture in a 650 mOsm kg H2O−1 medium in which glutamine was the sole nitrogen source. Colony formation was achieved in 6–7 weeks in a modified protoplast culture medium in which a reduction in the concentrations of both calcium and carbon was essential for sustained divisions. To maintain cell suspension growth, it was necessary to subculture every three weeks to a 170 mOsm kg H2O−1 medium. Lowering the C/N ratio did not support better growth. Phenolic compounds were detected in stationary phase cultures. Analysis by HPLC indicated that the cinnamate pathway was involved in their synthesis. After 3 and 7 months of culture, 65 and 74%, respectively, of protoplast-derived cells had a nuclear DNA content comparable to that of leaf protoplasts.Keywords
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