Interspecific somatic hybrids between a diploid potato clone DG 81-68 susceptible to Phytophthora infestans (Mont.) de Bary and a resistant diploid tuber-bearing species Solanum 9 michoacanum were generated and analyzed. About 30 regenerants displaying an intermediate morphology were obtained as a result of three separate PEG-mediated fusion experiments. The RAPD analysis confirmed the hybridity of all the regenerants. About 50% of the hybrid plants exhibited vigorous growth and were stable in culture, while the rest of them rooted poorly and grew slowly in vitro. Most of the hybrid clones were at the tetraploid level (70%), while 30% of the clones examined were at the hexaploid level. The S. 9 michoacanum (?) DG 81-68 hybrids with growth anomalies were aneuploid. The variation in late blight resistance of the hybrid clones was found in detached leaflet tests, with enhanced resistance characteristic for three tetraploid hybrids.
The interspecific somatic hybrids 4x S. villosum (+) 2x S. tuberosum clone DG 81-68 (VT hybrids) were obtained and characterized molecularly and cytogenetically. The morphology of fusion-derived plants was intermediate in relation to the parental species. The expected ploidy level of the regenerants was 6x for the VT hybrids, but the real ploidy of the hybrids varied, with some of them being euploids, and others - aneuploids. The hybridity of the regenerants was verified by random amplified polymorphic DNA (RAPD) analysis. Despite the variation in ploidy, the RAPD patterns of the hybrids were mostly uniform, suggesting similarity of the genotypes of the VT clones. Genomic in situ hybridization (GISH) analysis discriminated between the chromosomes of both parental genomes in VT somatic hybrids and also confirmed their hybridity. The resistance of VT somatic hybrids to Phytophthora infestans was evaluated and all of the hybrids proved to be highly resistant. In search of the mechanisms involved in resistance of the Solanum species to P. infestans, the biochemical reactions occurring early after elicitor treatment were studied. The production of reactive oxygen species (ROS), as one of the earliest reactions induced by pathogens or their elicitors, was examined in the resistant wild species S. villosum, susceptible S. tuberosum clone DG 81-68 and in the VT hybrid, resistant to P. infestans. After treatment of the leaves with elicitor, the relative increase in ROS production was higher in leaves of the susceptible potato clone than in the resistant plants of S. villosum and the somatic hybrid.
An attempt was made to change the proportion of the parental genomes in interspecific hybrids Solanum nigrum ? S. tuberosum (ngr ? tbr) by means of repeated protoplast fusion. In order to enlarge the potato input into the hybrid genome, the protoplasts of two ngr ? tbr hybrids of different ploidy (7x and 8x) were fused with the protoplasts of two different diploid potato clones in three combinations. Protoclonal variability was studied in three populations of new ngr ? tbr allopolyploids maintained in vitro. The absolute nuclear DNA content (2C) was measured using flow cytometry to estimate the ploidy of the hybrids. The ploidy level of the selected clones was verified by chromosome counts in root meristems. The newly synthesized allopolyploids (75 clones) showed only a small gain in nuclear DNA content above the mean value determined for the parents, instead of the expected addition of an entire diploid potato genome to the combined parental ngr ? tbr genome. An increase in nuclear DNA was observed mostly in the clones having the 7x hybrid as a parent (75% of allopolyploids from two combinations). When the 8x hybrid was used as a parent, only two allopolyploids (5%) exhibited a significantly increased nuclear DNA content. The 8x level of ngr ? tbr allopolyploids was shown to be stable and was only occasionally exceeded. Somatic hybrids ngr ? tbr offer a model system for studying the molecular mechanism(s) and processes involved in stabilization and establishment of the synthetic Solanum allopolyploids.
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