Experimental Hybridization between Some Marine Coenocytic Green Algae Using Protoplasms Extruded in vitro

A unique ability to form protoplasts in vitro from the extruded protoplasm has been found in a number of the marine coenocytic green algae (Tatewaki and Nagata 1970; Rietema 1973; Ishizawa and Wada 1979; Kim and Klotchkova 2004). After cell injury, extruded cell organelles in seawater aggregate to form compact protoplasmic masses; a process seems to be mediated by a lectin-carbohydrate complementary system (Kim et al. 2002; Klotchkova et al. 2003). The agglutinated protoplasmic masses soon change into spheres, i.e., protoplasts, surrounded with an envelope (Kobayashi and Kanaizuka 1985; Pak et al. 1991). Cytochemical and ultrastructural studies revealed that this envelope is not a lipid-based plasma membrane but is a membrane composed of some polysaccharides (Kim et al. 2001; Klotchkova et al. 2003). The envelope transforms into a plasma membrane within several hours, presumably by incorporation of disintegrated plasma membrane of the precursor cell from the interior of protoplast onto its surface (Kim et al. 2001; Klotchkova et al. 2003). The protoplasts develop a new cell wall in a day or two. In some species these cells grow into plants, whereas in other species they lose the ability to grow but remain as an enlarged single cell, producing numerous aplanospores or swarmers inside. It was therefore suggested that wound-induced protoplast formation in vitro may be a natural dispersal mechanism (Kim and Klotchkova 2004). Because coenocytic greens often grow in close proximity and their protoplasms may come into physical contact in nature, it is important to investigate if they can intermix while assembling the protoplast. The dissociated cell components of the coenocytic protoplasm, separated by forcing it through sucrose gradient solution by ultracentrifugation, could be reunited into live cells and thus the formation of new species by mixing cell components from different Algae Volume 20(3): 239-249, 2005