ABSTRACT
This chapter presents the techniques and approaches used in most studies of protoplast isolation and culture. The isolation of protoplasts from plant cells was first achieved by microsurgery on plasmolyzed cells. The plant species, the conditions of plant growth, plant age, protoplast isolation, and protoplast culture are often critical for sustained division of protoplasts. Callus and cell suspensions are frequently used for protoplast isolation and they offer aseptic conditions from the beginning. The combination of agarose plating and bead culture has been shown to dramatically improve plating efficiencies of protoplasts in many species tested. Generally, the basic constituents in the most frequently used protoplast media are similar to those used for cell cultures. A suitable carbon source is of great importance in a protoplast culture medium. The majority of protoplast culture media contain one or more auxins plus one or two cytokinins to stimulate protoplast division and growth. Protoplast isolation and culture is a well-established technique for many plant species.
