A new Method to Isolate Protoplasts from the Cyanobacterium Anabaena cylindrica

Abstract

Summary The present study was carried out to establish a new isolation method for cyanobacterial protoplasts. Conventional lysozyme treatment is appropriate for the isolation of spheroplasts from cyanobacteria, but not to produce protoplasts. Using Anabaena cylindrica, Mastigocladus laminosus and Oscillatoria princeps as materials, the effects of various enzyme treatments on protoplast isolation were examined. When Anabaena cylindrica was treated with N-acetylmuramidase SG, protoplasts lacking cell wall residues were obtained. The enzyme digested the layer II of the Anabaena cell wall, as with lysozyme. However, N-acetylmuramidase SG was effective to the digestion of layer IV, but not lysozyme. The treatments with lysozyme, achromopeptidase TBL-1, trypsin and ±-amylase were effective for the digestion of Anabaena cell wall respectively, but led to intracellular structure destruction, and moreover made phycocyanin flow out before the protoplasts were isolated. On the other hand, Anabaena protoplasts produced by N-acetylmuramidase SG treatment were intact in their fine structure and functionally active just after the isolation.