ABSTRACT
Plant protoplasts are dened as cells that have had their rigid cellulose cell wall removed without damaging the external cell membrane that surrounds the nucleus and cytoplasm. Under appropriate osmoticum conditions, protoplasts are perfectly spherical in shape. Initial efforts to isolate protoplasts began in the late 1800s using mechanical methods, but these met with limited success. The extraction of cellulase, macerase, and pectinase enzymes from various fungi during the mid-1900s provided new opportunities for protoplast isolation via enzymatic digestion, resulting in the landmark paper on plant protoplast isolation by Edward C. Cocking (Cocking, 1960), who used a crude cellulase preparation to isolate protoplasts from roots of tomato seedlings. Cell-wall digestion is generally carried out in a cocktail solution of enzymes and elevated osmoticum (for plasmolysis, usually sugars or sugar alcohols), empirically determined for any particular species/ explant. Protoplasts can now be routinely isolated on a large scale from the most important crops and ornamental species from nearly any plant part or cultured cells. Protoplasts have been used in many applications in plant biotechnology and molecular biology research, including somatic hybridization and cybridization, transformation, transient assays, protein-protein interactions, and screening for disease resistance. This chapter will discuss practical applications of evolving plant protoplast technologies.
