Abstract
Auxotroph complementation was achieved by pairwise protoplast fusion using three cell lines of Datura innoxia P. Mill., Pn1, IV-1 and JM3, having an obligate requirement for growth of pantothenate, isoleucine and valine, and threonine, respectively. Protoplast fusion was induced in a fusion solution containing polyethylene glycol (25%, MW 4000), glycine (0.05 M), and calcium chloride (0.1 M) at pH 8.5. After the fusion treatment, the protoplasts were cultured initially in a non-selective medium which was gradually diluted with minimal medium. Putative prototrophic hybrid colonies were rescued on selective feeder plates prepared with wild-type Datura cells and selective medium. The use of feeder plates was a necessary step to recover appreciable number of hybrid colonies. Somatic hybrids were identified on the basis of their growth in minimal medium, chromosome numbers and restoration of organogenetic capacity.
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Saxena, P.K., King, J. Complementation by intraspecific protoplast fusion of auxotrophic cell lines of Datura innoxia P. Mill.. Plant Cell Tiss Organ Cult 9, 61–71 (1987). https://doi.org/10.1007/BF00046080
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DOI: https://doi.org/10.1007/BF00046080