Summary
The Dutch potato cultivar Bintje has been transformed by Agrobacterium strain LBA1060KG, which contains two plasmids carrying three different DNAs (TL- and TR-DNA on the Agrobacterium rhizogenes plasmid and TKG-DNA on the pBI121 plasmid). Several transformed root clones were obtained after transformation of leaf, stem, and tuber segments, and plants were then regenerated from these root clones. The expression of the various marker genes [rol, opine, β-glucuronidase (GUS), and neomycin phosphotransferase (NPTII)] was determined in several root clones and in regenerated plants. The selection of vigorously growing root clones was as efficient as selection for kanamycin resistance. In spite of the location of NPTII and GUS genes on the same T-DNA, 17% of the root clones did not show GUS activity. Nevertheless, Southern blot analysis showed that these root clones contained at least three copies of the GUS gene. Sixty-four per cent of the root clones contained opines. The expression of these genes, however, was negatively correlated with plant regeneration capacity and normal plant development. The differential expression of the marker genes in the transgenic potato tissues is discussed.
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Anzai H, Yoneyama K, Yamaguchi I (1989) Transgenic tobacco resistant to a bacterial disease by the detoxification of a pathogenic toxin. Mol Gen Genet 219:492–494
Bradford M (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein dye binding. Anal Biochem 72:248–254
Cardarelli M, Spano L, Mariotti D, Mauro ML, Costantino P (1987) The role of auxin in hairy root induction. Mol Gen Genet 208:457–463
Comai L, Faccioti D, Hyatt WR, Thompson G, Rose RE, Stalker DM (1985) Expression in plants of a mutant aroA gene from Salmonella typhimurium confers tolerance to glyphosate. Nature 317:741–744
Dellaporta SL, Wood J, Hicks JB (1983) A plant DNA minipreparation: Version II. Plant Mol Biol Rep 1:19–24
De Vries-Uijtewaal E, Gilissen LJW, Flipse E, Sree Ramulu K, Stiekema WJ, De Groot B (1989) Fate of introduced genetic markers in transformed root clones and regenerated plants of monohaploid and diploid potato genotypes. Theor Appl Genet 78:185–193
Dynan WS, Tijan R (1985) Control of eukaryotic messenger RNA synthesis by sequence-specific DNA-binding proteins. Nature 316:774–778
Haas MJ, Dowding JE (1975) Aminoglycoside-modifying enzymes. In: Colowick SP, Kaplan NO (eds) Methods in enzymology XLIII. Academic Press, New York, pp 611–627
Hamill JD, Prescott A, Martin C (1987) Assessment of the efficiency of cotransformation of the T-DNA of disarmed binary vectors derived from Agrobacterium tumefaciens and the T-DNA of A. rhizogenes. Plant Mol Biol 9:573–584
Hänisch ten Cate ChH, Ramulu K Sree, Dijkhuis P, De Groot B (1987) Genetic stability of cultured hairy roots induced by Agrobacterium rhizogenes on tuber disks of potato cv. Bintje. Plant Sci 49:217–222
Hänisch ten Cate ChH, Ennik E, Roest S, Ramulu KS, Dijkhuis P, De Groot B (1988) Regeneration and characterization of plants from potato root lines transformed by Agrobacterium rhizogenes. Theor Appl Genet 75:452–459
Hänisch ten Cate ChH, Loonen AEHM, Ottaviani MP, Ennik L, Van Eldik G, Stiekema WJ (1990) Frequent spontaneous deletions of Ri T-DNA in Agrobacterium rhizogenes- transformed potato roots and regenerated plants. Plant Mol Biol 14:735–741
Hepburn AG, Clarke LE, Pearson L, White J (1983) The role of cytosine methylation in the control of nopaline synthase gene expression in a plant tumor. J Mol Appl Genet 2:315–339
Jefferson RA, Kavanagh TA, Bevan MT (1987) GUS fusions: β-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J 6:3901–3907
Jouanin L (1984) Restriction map of an agropine-type Ri plasmid and its homologies with Ti plasmids. Plasmid 12:91–102
Matzke MA, Primig M, Trnovsky J, Matzke AFM (1989) Reversible methylation and inactivation of marker genes in sequentially transformed tobacco plants. EMBO J 8:643–649
Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:473–497
Offringa IA, Melchers LS, Regensburg-Tuik AJG, Costantino P, Schilperoort RA, Hooykass PJJ (1986) Complementation of Agrobacterium tumefaciens tumor-inducing aux mutants by genes from the TR region of the Ri plasmid of Agrobacterium rhizogenes. Proc Natl Acad Sci USA 83:6935–6939
Ooms G, Twell D, Bossen ME, Hoge JHC, Burell MM (1986) Developmental regulation of Ri TL-DNA gene expression in roots, shoots, and tubers of transformed potato (Solanum tuberosum cv Desiree) Plant Mol Biol 6:321–330
Oono Y, Handa T, Kanaya K, Hchimiya H (1987) The TL-DNA gene of Ri plasmids responsible for dwarfness of tobacco plants. Jpn J Genet 62:501–505
Ottaviani MP, Schel J, Hänisch ten Cate ChH (1990) Variation in structure and plant regeneration of Agrobacterium rhizogenes-transformed and control roots of the potato cv Bintje. Plant Cell Tiss Org Cult 20:25–34
Petit A, David C, Dahl GA, Ellis JG, Guyon P, Casse-Delbart F, Tempé J (1983) Further extension of the opine concept: plasmids in Agrobacterium rhizogenes cooperate for opine degradation. Mol Gen Genet 190:204–214
Saedler H, Linn F, Meyer P (1989) Engineering of a new flower color variety of petunia. Horticultural Biotechnology Symposium, University of California, Davis/Ca, pp 22–23
Schmülling T, Schell J, Spena A (1988) Single genes from Agrobacterium rhizogenes influence plant development. EMBO J 7:2621–2629
Shah DM, Horsch RB, Klee HJ, Kishore GM, Winter JA, Tumer NE, Hironaka CM, Sanders PR, Gasser CS, Aykent S, Siegel NR, Rogers SG, Frayler RT (1986) Engineering herbicide tolerance in transgenic plants. Science 233:478–481
Shen WH, Petit A, Guern J, Tempé J (1988) Hairy roots are more sensitive to auxin than normal roots. Proc Natl Acad Sci USA 85:3417–3421
Simpson RB, Spielmann A, Margassion L, McKnight TD (1986) A disarmed binary vector from Agrobacterium tumefaciens functions in Agrobacterium rhizogenes: frequent Cotransformation of two distinct T-DNAs. Plant Mol Biol 6:403–415
Skoog F, Miller CO (1957) Chemical regulation of growth and organ formation in plant tissues cultured in vitro. Symp Soc Exp Biol 11:118–131
Spena A, Schmülling T, Koncz C, Schell JS (1987) Independent and synergistic activity of rol A, B, and C loci in stimulating abnormal growth in plants. EMBO J 6:3891–3899
Stiekema WJ, Heidekamp F, Louwerse JD, Verhoeven HA, Dijkhuis P (1988) Introduction of foreign genes into potato cultivars Bintje and Desiree using an Agrobacterium tumefaciens binary vector. Plant Cell Rep 7:47–50
Sukhapinda K, Spivey R, Simpson RB, Shahin EL (1987) Transgenic tomato (Lycopersicon esculutum L.) transformed with a binary vector in Agrobacterium rhizogenes: non-chimeric origin of callus clone and low copy numbers of integrated vector of T-DNA. Mol Gen Genet 206:491–497
Taylor BH, Amasino RM, White FF, Nester EW, Gordon MP (1985) T-DNA analysis of plants regenerated from hairy root tumors. Mol Gen Genet 201:554–557
Tepfer D (1983) The potential uses of Agrobacterium rhizogenes in the genetic engineering of higher plants: nature got there first. In: Lurquin PF, Kleinhofs A (eds) Genetic engineering in eukaryotes. Plenum Press, New York, pp 153–164
Vilaine F, Casse-Delbart F (1987) Independent induction of transformed roots by the TL and TR regions of the Ri plasmid of agropine-type Agrobacterium rhizogenes. Mol Gen Genet 206:17–23
Visser RGF, Jacobsen E, Witholt B, Feenstra WJ (1989) Efficient transformation of potato (Solanum tuberosum) with a binary vector in Agrobacterium rhizogenes. Theor Appl Genet 78:594–600
White FF, Taylor BH, Huffman GA, Gordon MP, Nester EW (1985) Molecular and genetic analysis of the root-inducing plasmid of Agrobacterium rhizogenes: morphogenic loci are divided between two regions of transferred DNA. J Bacteriol 164:33–44
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Communicated by Yu. Gleba
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Ottaviani, M.P., Hänisch ten Cate, C.H. Cotransformation and differential expression of introduced genes into potato (Solanum tuberosum L.) cv Bintje. Theoret. Appl. Genetics 81, 761–768 (1991). https://doi.org/10.1007/BF00224987
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DOI: https://doi.org/10.1007/BF00224987