Somatic hybridization by microfusion of defined protoplast pairs in Nicotiana: morphological, genetic, and molecular characterization

Spangenberg, G.; Osusky, M.; Oliveira, M.M.; Freydl, E.; Nagel, J.; Pais, M.S.; Potrykus, I.

Tag. Theoretical and Applied Genetics. Theoretische und Angewandte Genetik 80(5): 577-587

1990


ISSN/ISBN: 0040-5752
PMID: 24221062
DOI: 10.1007/bf00224215
Accession: 007803639

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Abstract
Somatic hybrid/cybrid plants were obtained by microfusion of defined protoplast pairs from malefertile, streptomycin-resistant Nicotiana tabacum and cytoplasmic male-sterile (cms), streptomycin-sensitive N. tabacum cms (N. bigelovii) after microculture of recovered fusants. Genetic and molecular characterization of the organelle composition of 30 somatic hybrid/cybrid plants was performed. The fate of chloroplasts was assessed by an in vivo assay for streptomycin resistance/ sensitivity using leaf explants (R0 generation and R1 seedlings). For the analysis of the mitochondrial (mt) DNA, species-specific patterns were generated by Southern hybridization of restriction endonuclease digests of total DNA and mtDNA, with three DNA probes of N. sylvestris mitochondrial origin. In addition, detailed histological and scanning electron microscopy studies on flower ontogeny were performed for representative somatic hybrids/cybrids showing interesting flower morphology. The present study demonstrates that electrofusion of individually selected pairs of protoplasts (microfusion) can be used for the controlled somatic hybridization of higher plants.

Somatic hybridization by microfusion of defined protoplast pairs in Nicotiana: morphological, genetic, and molecular characterization