Transformation of Plant Protoplasts in Vitro

Abstract

In genetic manipulation one can distinguish between the mixing of more or less complete genomes and the more accurate introduction of well defined genes. In the first case use is being made of somatic cell fusion and the uptake of isolated chromosomes and organelles. It might particularly contribute in the transfer of complex genetic traits, like polygenic complexes, that affect yield in agricultural production. In the second case use is being made of recombinant DNA and transformation of protoplasts. By this approach only one or a small number of genes are transferred into an otherwise unaltered genetic background, which has many advantages over less accurate procedures. It is quite clear that plant genetic manipulation is at an early stage of development and that much has still to be learned. Nevertheless, it is proven that this technology can overcome restrictions on gene flow between widely different organisms. Its use will enable us to understand e.g. genomic organization and regulation of gene expression in higher plants. Unless much more is known about this subject and the molecular processes, that underly plant phenotype in general, it can not be expected that somatic and molecular genetics will significantly contribute to applications of practical use. Here, we only deal with the transformation of plant cells, which requires a procedure for introducing DNA into cells followed by its expression. The major obstacle to DNA uptake in plant cells is the cell wall, but this can be circumvented by using plant protoplasts, i.e. cells freed of their cell walls by enzymatic digestion. Using appropriate media, protoplasts regenerate a new cell wall and subsequently divide. After sustained cell divisions small cell clumps arise.