Abstract
During culture, Brassica napus leaf protoplasts become swollen and finally degrade. Both morphological and biochemical approaches revealed that the process through which the death occurred were subjected to apoptotic-like manner. The Protease activity remained lower in leaves, but gradually increased during the culture. Several protease inhibitors added to the enzyme solution for protoplast isolation had no apparent effect on protoplast survival. The previous results showed that the amount of extracellular hydrogen peroxide and superoxide present in the medium during the isolation of leaf protoplasts may not be involved in triggering apoptosis from B. napus. DCFH-DA staining visualizes intracellular active oxygen species (AOS), especially hydrogen peroxide. Our data showed that greater levels of AOS were accumulated in B. napus chloroplasts. However, addition of radical scavengers to both the isolation and the culture medium had no effect on the cell death of B. napus protoplasts.
