Transformation and regeneration of orchardgrass protoplasts
The idea of using embryogenic suspension cultures as a source material for protoplasts was first popularized by I.K. Vasil and co-workers. The reasons for the success of embryogenic suspension cells giving totipotent protoplasts are, in hindsight, very clear. First, the cells in these suspension cultures (indeed, in all suspension cultures) are dividing relatively rapidly. The resultant protoplasts are already programmed to divide and, in fact, do so after only three to four days following plating. Second, as embryogenic cultures, every cell in every aggregate should have the ability to regenerate into somatic embryos which can convert into plantlets. Since non-embryogenic cultures rarely are able to produce plantlets, it seems unlikely such cultures could give protoplasts capable of doing much more than reconstituting a cell wall and going through a division or two. Starting with meristems of some type as the protoplast source material would seem to satisfy the above criteria but they are hard to isolate en masse and actually contain only a few percent of cells dividing. This latter point is also true of embryogenic callus as the plating efficiency is always extremely poor with embryogenic callus-derived protoplasts. Thus, currently the only source from which to reliably obtain large quantities of totipotent monocotyledonous protoplasts is from embryogenic suspension cultures.
KeywordsSomatic Embryo Suspension Culture Protoplast Isolation Embryogenic Suspension Cell Embryogenic Suspension Culture
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