| Abstract | This chapter discusses the plant protoplast culture. Protoplasts have been isolated from both intact plants and cell cultures of many plant species. Leaves are an especially suitable cell source because mesophyll cells have relatively thin walls which are readily degraded. It may be necessary to remove the epidermis because enzymes have difficulty in penetrating the cuticle. Pretreatment with α-glucuronidase or pectinglucosidase can partially substitute for epidermis stripping. Other plant parts such as root tips and embryos may also be used but generally have cells with more resistant walls. Rapidly growing cell cultures are a most suitable source of protoplasts. Old cells or cells from slowly growing cultures often have walls which are highly resistant to enzyme degradation. The medium should be slightly hypertonic because it appears that plasmolysis hastens the wall degradation, perhaps by allowing enzyme attack from the inner surface of the wall. Protoplasts appear to be more stable than the cells in the incubation mixtures. Generally, all cells not converted to protoplasts are dead after 18 hours incubation with the enzyme mixture. |
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