Abstract
Leaf protoplasts of two wild species, Solanum nigrum var. gigantea (S. ngr gig) and S. bulbocastanum Dun. (S. blb), were electrofused with leaf protoplasts of two diploid potato clones, H-8105 and ZEL-1136, respectively, in order to confer the late blight-resistance from the wild species to the cultivated potato. The S. ngr gig mesophyll (+) H-8105 mesophyll combination resulted in regenerants of mostly normal ngr phenotype. Two regenerants from this combination were proved to be true hybrids by RAPD analysis but they rooted poorely in vitro and did not survive the transfer to soil. The S. ngr gig (+) H-8105 fusion combination was also performed with H-8105 cell suspension derived protoplasts enabling an easy identification of interspecific fusants on basis of their intermediate morphology. From the S. ngr gig mesophyll (+) H-8105 cultured cell combination, many abnormal shoots were regenerated. The two lines which survived had normal ngr phenotype but the presence of tuberosum (tbr) genome in those regenerants was not confirmed by RAPD analysis. No plants with tbr phenotype were obtained from both of S. ngr gig (+) H-8105 combinations. On the contrary, when S. blb mesophyll protoplasts were electrofused with ZEL-1136 mesophyll protoplasts, all regenerated plants had tbr phenotype, indicating much lower morphogenetic potential of S. bulbocastanum in comparison with that of S. nigrum var. gigantea. However, the hybridity of those regenerants has not been confirmed by RAPD analysis with two different primers. The efficiency of the applied fusion procedure and analysis of the regenerants is discussed.
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Austin S., Baer M., Ehlenfeldt M., Kazmierczak P.J., Helgeson J.P. 1985. Intra-specific fusions in Solanum tuberosum. Theor. Appl. Genet., 71: 172–175.
Austin S., Pohlman J.D., Brown C.R., Mojtahedi H., Santo G.S., Douches D.S., Helgeson J.P. 1993. Interspecific somatic hybridization between Solanum tuberosum L. and S. bulbocastanum Dun. as a means of transferring nematode resistance. Am. Potato J., 70: 485–495.
Debnath S.C., Wenzel G. 1987. Selection of somatic fusion products in potato by hybrid vigour. Potato Res., 30: 371–380.
Fish N., Karp A., Jones M.G.K. 1988. Production of somatic hybrids by electrofusion in Solanum. Theor. Appl. Genet., 76; 260–266.
Fish N., Steele S.H., Jones M.G.K. 1988. Field assessment of dihaploid Solanum tuberosum and S. brevidens somatic hybrids. Theor. Appl. Genet., 76: 880–886.
Gamborg O.L., Miller R.A., Ojima K. 1968. Nutrient requirements of suspension cultures of soybean root cells. Exp. Cell Res., 50: 151–158.
Haberlach G.T., Cohen B.A., Baer M.A., Towill L.E. Helgeson J.P. 1985. Isolation, culture and regeneration of protoplasts from potato and several related Solanum species. Plant Sci., 39: 67–74.
Helgeson J.P., Hunt G.J., Haberlach G.T., Austin S. 1986. Somatic hybrids between Solanum brevidens and Solanum tuberosum: Expression of a late blight resistance gene and potato leaf roll resistance. Plant Cell Rep., 3: 212–214.
Helgeson J.P., Pohlman J.D., Austin S., Haberlach G.T., Wielgus S.M., Ronis D., Zambolim L., Tooley P., McGrath J.M., James R.M., Stevenson W.K. 1998. Somatic hybrids between Solanum bulbocastanum and potato: a new source of resistance to late blight. Theor. Appl. Genet., 96: 738–742.
Horsman K., Bergervoet J.E.M., Jacobsen E. 1997. Somatic hybridization between Solanum tuberosum and species of the S. nigrum complex: Selection of vigorously growing and flowering plants. Euphytica, 96: 345–352.
Jones M.G.K. 1988. Fuzing plant protoplasts. Trends Biotechnol., 6: 153–158.
Karp A., Nelson R.S., Thomas E., Bright S.W.J. 1982. Chromosome variation in protoplast-derived potato plants. Theor. Appl. Genet., 63: 1–7.
Laferriere L. T., Helgeson J. P., Allen C. 1999. Fertile Solanum tuberosum + S. commersonii somatic hybrids as sources of resistance to bacterial wilt caused by Ralstonia solanacearum. Theor. Appl. Genet. 98: 1272–1278.
Maciejewska U., Skierski J. S., Szczerbakowa A. 1999. Nuclear DNA content of Solanum species grown in vitro, as determined by flow cytometry. Acta Physiol. Plant., 21: 37–43.
Mattheij W. M., Eijlander R., de Konig J.R.A., Louwes K.M. 1992. Interspecific hybridization between the cultivated potato Solanum tuberosum subspecies tuberosum L. and the wild species S. circaeifolium subsp. circaeifolium Bitter exhibiting resistance to Phytophthora infestans (Mont.) de Bary and Globodera pallida (Stone) Behrens 1. Somatic hybrids. Theor. Appl. Genet., 83: 459–466.
Menczel L., Nagy F., Kiss Zs.R., Maliga P. 1981. Streptomycin resistant and sensitive somatic hybrids of Nicotiana tabacum + Nicotiana knightiana: Correlation of resistance to N. tabacum plastids. Theor. Appl. Genet., 59: 191–195.
Murashige T., Skoog F. 1962. A revised medium for rapid growth and bioassays with tobacco tissue culture. Physiol. Plant., 15: 473–497.
Nagy J.I., Maliga P. 1976. Callus induction and plant regeneration from mesophyll protoplasts of Nicotiana sylvestris. Z. Pflanzenphysiol., 78: 453–455.
Novy R. G., Helgeson J. P. 1994. Resistance to potato virus Y in somatic hybrids between Solanum etuberosum and S. tuberosum x S. berthaultii hybrid. Theor. Appl. Genet. 89: 783–786.
Oberwalder B., Ruoß B., Schilde-Rentschler L., Hemleben V., Ninnemann H. 1997. Asymmetric fusion between wild and cultivated species of potato (Solanum spp.) — detection of asymmetric hybrids and genome elimination. Theor. Appl. Genet., 94: 1104–1112.
Pawełczak A. 1991. Inbred selection of potatoes at the diploidal level. IHAR Bulletin, 177: 47–56 (in Polish).
Pawłowski P. 1995. Deformation of cells [Neurospora crassa slime] in an external periodic electric field. Ph.D. Thesis, Institute of Biochemistry and Biophysics PAS, Warsaw (in Polish).
Pehu E., Karp A., Moore K., Steele S., Dunckley R., Jones M.G.K. 1989. Molecular, cytogenetic and morphological characterization of somatic hybrids of dihaploid Solanum tuberosum and diploid S. brevidens. Theor. Appl. Genet., 78: 696–704.
Ramulu K.S., Dijkhuis P., Roest S. 1983. Phenotypic variation and ploidy level of plants regenerated from protoplasts of tetraploid potato (Solanum tuberosum L. cv. ‘Bintje’). Theor. Appl. Genet., 65: 329–338.
Rokka V.M., Xu Y.S., Kankila J., Kuusela A., Pulli S., Pehu E. 1994. Identification of somatic hybrids of dihaploid Solanum tuberosum lines and S. brevidens by species specific RAPD patterns and assessment of disease resistance of the hybrids. Euphytica, 80: 207–217.
Sambrook J., Fritsch E.F., Maniatis T. 1989. Molecular cloning. A laboratory manual, 2nd edition, V. 3, CSH Laboratory Press.
Sihachakr D., Haicour R., Chaput M.H., Barrientos E., Ducreux G., Rossignol L. 1989. Somatic hybrid plants produced by electrofusion between Solanum melongena L. and Solanum torvum Sw. Theor. Appl. Genet., 77: 1–6.
Sihachakr D., Haicour R., Serraf I., Barrientos E., Herbreteau C., Ducreux G., Rossignol L., Souvannavong V. 1988. Electrofusion for the production of somatic hybrid plants of Solanum melongena L. and Solanum khasianum C.B. Clark. Plant Sci., 57: 215–223.
Szczerbakowa A., Borkowska M., Wielgat B. 2000. Plant regeneration from the protoplasts of Solanum tuberosum, S. nigrum and S. bulbocastanum. Acta Physiol. Plant. 22: 3–10.
Tempelaar M.J., Duyst A., de Vlas S.Y., Krol G., Symonds C., Jones M.G.K. 1987. Modulation and direction of the electrofusion response in plant protoplasts. Plant Sci., 48: 99–105.
Tempelaar M.J., Jones M.G.K. 1985. Fusion characteristics of plant protoplasts in electric fields. Planta, 165: 205–216.
Thieme R., Darsow U., Gavrilenko T., Dorokhov D., Tiemann H. 1997. Production of somatic hybrids between S. tuberosum L. and late blight resistant Mexican wild potato species. Euphytica, 97: 189–200.
Waara S., Pijnacker L., Ferwerda M.A., Wallin A., Eriksson T. 1992. A cytogenetic and phenotypic characterization of somatic hybrid plants obtained after fusion of two different dihaploid clones of potato (Solanum tuberosum L.). Theor. Appl. Genet., 85: 470–479.
Wenzel G., Schieder O., Przewozny T., Sopory S.K., Melchers G. 1979. Comparison of single cell culture derived Solanum tuberosum L. plants and a model for their application in breeding programs. Theor. Appl. Genet., 55: 49–55.
Xu Y.S., Clark M.S., Pehu E. 1993. Use of RAPD markers to screen somatic hybrids between Solanum tuberosum and S. brevidens. Plant Cell Rep., 12: 107–109.
Yamada T., Misoo S., Ishii T., Ito Y., Takaoka K., Kamijima O. 1997. Characterization of somatic hybrids between tetraploid Solanum tuberosum L. and dihaploid S. acaule. Breeding Sci., 47: 229–236
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Szczerbakowa, A., Maciejewska, U., Pawłowski, P. et al. Electrofusion of protoplasts from Solanum tuberosum, S. nigrum and S. bulbocastanum . Acta Physiol Plant 23, 169–179 (2001). https://doi.org/10.1007/s11738-001-0005-1
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DOI: https://doi.org/10.1007/s11738-001-0005-1