Monday, 14 November 2016

Selection of Marker-Free Transgenic Plants Using the Oncogenes (ipt, rol A, B, C) of Agrobacterium as Selectable Markers

Published Date
Volume 66 of the series Forestry Sciences pp 25-46

  • Hiroyasu Ebinuma
  • Koichi Sugita
  • Etsuko Matunaga
  • Saori Endo
  • Takehide Kasahara


Recently, it has become possible to modify the genetic traits of a wide variety of plants using genetic engineering. The conventional breeding of forest trees requires long periods of time to modify specific traits due to their long generation and rotation cycles. Genetic engineering has great potential for overcoming these hurdles by incorporating the desired genes into woody plants. In both hardwoods and conifers, genetic modification by DNA transfer has been reported in many species including poplar, sweetgum, eucalyptus, larch, spruce and pine (review: Ahuja, 1987, 1991; Sederoff et al., 1987; Ahuja et al., 1996). However, such genetic modification has still been difficult to achieve with many important woody plant species using the current transformation methods, because the regeneration frequency of woody plants is very low when compared with that of herbaceous plants. Therefore, we need to improve the transformation methods to facilitate genetic engineering in a wide variety of woody plants.


  1. Ahuja, M. R. (1987) Gene transfer in forest trees, in. J. W. Hanover and D. E. Keathley (eds.) Genetic manipulation of woody plants, Plenum Press, New York, pp. 24–41.
  2. Ahuja, M. R. (eds.) (1991) Woody plant biotechnology, Plenum Press, New York, p373.
  3. Ahuja, M. R., W. Boerjan and D. B. Neal (eds.) (1996) Somatic cell genetics and molecular genetics of trees, Kluwer Academic Publishers, Dordrecht, The Netherlands, p287.CrossRef
  4. Akiyoshi, D. E., H. Klee, R. M. Amasino, E. W. Nester and M. P. Gordon (1984) T-DNA of Agrobacterium tumefaciens encode an enzyme of cytokinin biosynthesis, Proc. Natl. Acad. Sci. USA81, 5994–5998.PubMedCrossRef
  5. Araki, H., A. Jearnpipatkul, H. Tatsumi, T. Sakurai, K. Ushino, T. Muta and Y. Oshima (1987) Molecular and functional organization of yeast plasmid pSRI, J. Mol. Biol182, 191–203.CrossRef
  6. Barry, G. F., S. G. Rogers, R.T. Fraley, and L. Brand (1984) Identification of a cloned cytokinin biosynthetic gene, Proc. Natl. Acad. Sci. USA81, 4776–4780.PubMedCrossRef
  7. Belzile, F., M. W. Lassner, Y. Tong, R. Khush, and J. 1. Yoder (1989) Sexual transmission of transposed Activatorelements in transgenic tomatoes, Genetics 123, 181–189.PubMed
  8. Brzobohatÿ, B., I. Moore and K. Palme (1994) Cytokinin metabolism: implication of plant growth and development, Plant Mol. Biol26, 1483–1497.PubMedCrossRef
  9. Christey M. C. (1997) Transgenic crop plants using Agrobacterium rhizogenes-mediated transformation, in. P. M. Doran (eds.) Hairy roots: culture and application, Hardwood Academic Publishers, Amsterdam, The Netherlands, pp. 99–111.
  10. Christou, P. (1996) Transformation technology, Trends in Plant Science 1, 423–431.CrossRef
  11. Cui, M., T. Handa and K. Takayanagi (1998) Transformation ofAntirrhinum majus L. by MAT vector, Japan J. Breed48 (Suppl. 2), 305.
  12. Dale, E. C. and D. W. Ow (1991) Gene transfer with subsequent removal of the selection gene from the host genome, Proc. Natl. Acad. Sci. USA88, 10558–10562.PubMedCrossRef
  13. Ebinuma, H., K. Sugita, E. Matsunaga and M. Yamakado (1997a) Selection of marker-free transgenic plants using the isopentenyl transferase gene as a selectable marker, Proc. Natl. Acad. Sci. USA 94, 2117–2121.PubMedCrossRef
  14. Ebinuma, H., K. Sugita, E. Matsunaga and M. Yamakado (1997b) Principle of MAT vector, Plant Biotech14, 133–139.CrossRef
  15. Endo, S., T. Kasahara, H. Ebinuma (1999) The improvement in selectable marker gene of MAT vector system, Japan J. Breed1 (Suppl. 1), 122.
  16. Estruch, J. J., E. Prinsen, H. V. Onckelen, J. Shell and A. Spena (1991) Viviparous leaves produced by somatic activation of an inactive cytokinin-synthesizing gene, Science 254, 1364–1367.PubMedCrossRef
  17. Fedoroff, N. (1989) Maize transposable elements, In D. E. Berg and M. M. Howe (eds.) Mobile DNAAm. Soc. Microbiol., Washington, DC, pp. 375–411.
  18. Gaudin, V., T. Vrain and L. Jouanin (1994) Bacterial genes modifying hormonal balances in plants, Plant Physiol. Biochem32, 11–29.
  19. Han, K. H., D. E. Keathley, J. M. Davis and M. P. Gordon (1993) Regeneration of a transgenic woody legume (Robinia pseudoacacia L., black locust) and morophorogical alterations induced by Agrobacterium rhizogenes-mediated transformation, Plant Sci88, 149–157.CrossRef
  20. Holt, D. C., V. J. Lay, E. D. Clarke, A. Dinsmore, I. Jepson, S. W. J. Bright and A. J. Greenland (1995) Characterization of the safener-induced glutathione S- transferase isoform II from maize, Planta 196, 295–302.PubMedCrossRef
  21. Hooykaas, P. J. J. and R. A. Schilperoort (1992) Agrobacterium and plant genetic engineering, Plant Mol. Biol19, 15–38.CrossRef
  22. James, D. J., A. J. Passey and D. J., Barbara (1990) Agrobacterium-mediated transformation of the cultivated strawberry (Fragaria xAnannassa Duch.) using disarmed binary vectors, Plant Sci69, 79–94.
  23. Kasahara, T. S. Endo, K. Sugita and H. Ebinuma (in preparation) Characterization of the ipt gene from Agrobacterium tumefaciens PO22 and of transgenic tobacco containing the ipt gene under control of native promoter.
  24. Kiyokawa, S. and K. Kobayashi, Y. Kikuchi, H. Kamada and H. Harada (1994) Root-inducing region of mikimopine type Ri plasmid pRi724, Plant Physiol104, 801–802.
  25. Lambert, C. and D. Tepfer (1992) Use of Agrobacterium rhizogenes to create transgenic apple trees having an altered organogenic response to hormones, Theor. Appl. Genet85, 105–109.CrossRef
  26. Li, Y., G. Hagen and T. J. Guilfoyle (1992) Altered morphology in transgenic tobacco plants that overproduce cytokinins in specific tissues and organs, Dev. Biol153, 386–395.
  27. Matsunaga, E., K. Sugita, T. Kasahara and H. Ebinuma (in preparation) High-efficiency transformation of hybrid aspen by using the chimeric ipt gene with a rbcS 3B promoter as a selectable marker.
  28. Medford, J. I., R. Horgan, Z. El-Sawi and H. J.Klee (1989) Alterations of endogenous cytokinins in transgenic plants using a chimeric isopentenyl transferase gene, Plant Cell 1, 403–413.PubMed
  29. Müller-Neumann, M., J. I. Yoder and P. Starlinger (1984) The DNA sequence of the transposable elements Ac of Zea mays LMol. Gen. Genet198, 19–24.CrossRef
  30. Nakano, M., Y. Hoshino and M. Mii (1994) Regeneration of transgenic plants of grapevine (Vitis vinifera L.) via Agrobacterium rhizogenes-mediated transformation of embryogenic calli, J. Exp. Bot45, 649–656.CrossRef
  31. Onouchi, H., K. Yokoi, C. Machida, H. Matsuzaki, Y. Oshima, K. Matsuoka, K. Nakamura and Y. Machida (1991) Operation of an efficient site-specific recombination system of Zygosaccharomyces rouxii in tobacco cells, Nucl. Acids Res19, 6373–6378.PubMedCrossRef
  32. Ooms, G., A. Kaup and J. Roberts (1983) From tumour to tuber; tumour cell characteristics and chromosome numbers of crown gall-derived tetraploid potato plants (Solanum tuberosum cv. ‘Maris Bard’), Theor. Appl. Genet66, 169–172.CrossRef
  33. Pythoud, F., V. P. Sinkar, E. W. Nester and M. P. Gordon (1987) Increased virulence of Agrobacterium rhizogenesconferred by the vir region of pTiBo542: application to genetic engineering of poplar, Bio/Technol5, 1323–1327.CrossRef
  34. Russell S. H., J. L. Hoopes and J. L. Odell (1992) Directed excision of a transgene from the plant genome, Mol. Gen. Genet234, 49–59.PubMed
  35. Schmulling, T., J. Beinsberger, J. D. Greef, J. Schell, H. V. Onckelen and A. Spena (1989) Construction of a heat-inducible chimeric gene to increase the cytokinin content in transgenic plant tissue, FEBS Letters 249, 401–406.CrossRef
  36. Schwartzenberg, K. V., P. Doumas, L. Jouanin and G. Pilate (1994) Enhancement of the endogenous cytokinin concentration in poplar by transformation with Agrobacterium T-DNA gene iptTree Physiol14, 27–35.CrossRef
  37. Sederoff, R., A. Stomp, B. Gwynn, E. Ford and C. Loopstra (1987) Application of recombinant DNA techniques to pines: A molecular approach to genetic engineering in forestry, in J. M. Bonga and D. J. Durzan (eds.) Cell and Tissue Culture in Forestry, Martinus Nijhoff Publishers, Dordrecht, The Netherlands, pp. 314–329.
  38. Shin, D. I., G. K. Podila, Y. Huang and D. E Karnosky (1994) Transgenic larch expressing genes for herbicide and insect resistance, Can. J. For. Res24, 2059–2067.CrossRef
  39. Slightom, J. L., M. Durand-Tardif, L. Jouanin and D. Tepfer (1986) Nucleotide sequence analysis of the TL-DNA ofAgrobacterium rhizogenes type plasmid, J. Biol261, 108–121.
  40. Smart, M. C., S. R. Scofield, M. W. Bevan and T. A. Dyer (1991) Delayed leaf senescence in tobacco plants transformed with tmr, a gene for cytokinin production in AgrobacteriumPlant Cell 3, 647–656.PubMed
  41. Smigocki, A. C. and F. A. Hammerschlag (1991) Regeneration of plants from peach embryo cells infected with a shooty mutant strain ofAgrobacteriumJ. Am. Soc. Hortic. Sci116, 1092–1097.
  42. Smigocki, A. C. and L. D. Owens (1988) Cytokinin gene fused with a strong promoter enhances shoot organogenesis and zeatin levels in transformed plant cells, Proc. Natl. Acad. Sci. USA85, 5131–5135.PubMedCrossRef
  43. Smigocki, A. C. and L. D. Owens (1989) Cytokinin-to-auxin ratios and morphology of shoots and tissues transformed by a chimeric isopentenyl transferase gene, Plant Physiol91, 808–811.PubMedCrossRef
  44. Sugita, K., E. Matsunaga and H. Ebinuma (in press) Effective selection system for generating marker-free transgenic plants independent of sexual crossing, Plant Cell Rep.
  45. Sugita, K., E. Matsunaga, T. Kasahara and H. Ebinuma (1999) The multiple gene introductions into plants using the MAT-vector, Japan J. Breed1 (Suppl. 1), 121.
  46. Sugita, M. and W. Gruissem (1987) Developmental, organ-specific, and light-dependent expression of the tomato ribulose-1,5-bisphosphate carboxylase small subunit gene family, Proc. Natl. Acad. Sci. USA84, 71047108.
  47. Takayama, Y (1968) Studies on the breeding of aspens (I): Height growth in the early stage of F1 seedlings of Populus sieboldii Miq. x P. grandidentata Michx, Nichirinshi 50, 267–273.
  48. Tepfer, D (1984) Transformation of several species of higher plants by Agrobacterium rhizogenes: sexual transmission of the transfomed genotype and phenotype, Cell 37, 959–967.PubMedCrossRef
  49. Thomashow, L. S., S. Reeves and M. E Thomashow (1984) Crown gall oncogenesis: evidence that a T-DNA gene from the Agrobacterium Ti plasmid pTiA6 encodes an enzyme that catalyses synthesis of indoleacetic acid, Proc. Natl. Acad. Sci. USA81, 5071–5075.PubMedCrossRef
  50. Vahala, T., T. Eriksson, E. Tillberg and B. Nicander (1993) Expression of a cytokinin synthesis gene from Agrobacterium tumefaciens T-DNA in basket willow (Salix viminalis)Physiologia Plantarum 88, 439–445.CrossRef
  51. Wabiko, H., M. Kagaya, I. Kodama, K. Masuda, Y. Kodama, H. Yamamoto, Y. Shibano and H. Sano (1989) Isolation and characterization of diverse nopaline type Ti plasmids ofAgrobacterium tumefaciens from Japan, Arch Microbiol152, 119–124.CrossRef
  52. White F. F., B. H. Taylor, G. A. Huffman and M. P. Gordon (1985) Molecular and genetic analysis of the transferred DNA regions of the root-inducing plasmid of Agrobacterium rhizogenesJ. Bacteriol164, 33–44.PubMed
  53. Yazawa, M., C. Suginuma, K. Ichikawa, H. Kamada and T. Akihama (1995) Regeneration of transgenic plants from hairy root of kiwi fruit (Actinidia deliciosa) induced by Agrobacterium rhizogenesBreeding Sci45, 241–244.
  54. Yoder, J. I. and A. P. Goldsbrough (1994) Transformation systems for generating marker-free transgenic plants, Bio/lechnol12, 263–267.CrossRef

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