Abstract
Apple (Malus × domestica) is one of the most consumed fruit crops in the world. The major production areas are the temperate regions, however, because of its excellent storage capacity it is transported to distant markets covering the four corners of the earth. Transformation is a key to sustaining this demand—permitting the potential enhancement of existing cultivars as well as to investigate the development of new cultivars resistant to pest, disease, and storage problems that occur in the major production areas. In this paper we describe an efficient Agrobacterium tumefaciens-mediated transformation protocol that utilizes leaf tissues from in vitro grown plants. Shoot regeneration is selected with kanamycin using the selectable kanamycin phosphotransferase (APH(3)II) gene and the resulting transformants confirmed using the scorable uidA gene encoding the bacterial β-glucuronidase (GUS) enzyme via histochemical staining. Transformed shoots are propagated, rooted to create transgenic plants that are then introduced into soil, acclimatized and transferred to the greenhouse from where they are taken out into the orchard for field-testing.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Zambryski, P., Temple, J., and Schell, J. (1989) Transfer and function of T-DNA genes from Agrobacterium Ti-and Ri-plasmids in plants. Cell 56, 193–201.
James, D.J., Passey, A.J., and Ruggini E. (1988) Factors affecting high frequency plant regeneration from apple leaf tissue cultured in vitro. J. Plant Physiol. 132, 148–154.
Fasolo, F., Zimmerman, R.H., and Fordham, I. (1989) Adventitious shoot formation on excised leaves of in vitro grown shoots of apple cultivar. Plant Cell Tissue Org. Cul. 16, 75–87.
Predieri, S., Fasolo, F., and Lalavasi, F. (1989) High frequency shoot regeneration from leaves of apple rootstock M.26 (Malus pumila Mill.). Plant Cell Tissue Org. Cult. 17, 133–142.
Welander, M. (1988) Plant regeneration from leaf and stem segments of shoots raised in vitro from mature apple trees. J. Plant Physiol. 132, 738–744.
Bevan, M.W. (1984) Binary Agrobacterium vectors for plants transformation. Nucleic Acids Res. 12, 8711–8721.
McBride, K.E. and Summerfelt, K.R. (1990) Improved binary vectors for Agrobacterium-mediated plant transformation. Plant Mol. Biol. 14, 269–276.
Wen-jun, S. and Forde, B. (1989) Efficient transformation of Agrobacterium spp. by high voltage electroporation. Nucleic Acids Res. 17, 8385.
James, D.J. and Dandekar, A.M. (1991) Regeneration and transformation of apple (Malus pumila Mill.). Plant Tissue Culture Manual: Fundamentals and Applications (Lindsey, K., ed.), Kluwer Academic Publishers, Dorderecht, The Netherlands, vol. 88, pp. 1–18.
James, D.J. and Thurbon, I.J. (1979) Rapid in vitro rooting of apple rootstock M.9. J. Hort. Sci. 54, 309–311.
Horsch, R.B., Fry, J.E., Hoffman, N.L., Eicholtz, D., Rogers, S.G., and Fraley, R.T. (1985) A simple and general method of transferring genes into plants. Science 227, 1229–1231.
James, D.J., Uratsu, S., Cheng, J., Negri, P., Viss, P., and Dandekar, A.M. (1993) Acetosyringone and osmoprotectants like betaine or proline synergistically enhance Agrobacterium mediated transformation of apple. Plant Cell Rep. 12, 559–563.
Hood, E.A., Helmer, G.L., Fraley, R.T., and Chilton, M-D. (1986) The hypervirulence of Agrobacterium tumefaciens A281 is encoded in a region of pTiBo542 outside the T-DNA. J. Bacteriol. 168, 1291–1301.
Van Larebeke, N., Engler, G., Holsters, M., Van den Elsacker, Zaenen, I., Schilperoort, R.A., and Schell, J. (1974) Large plasmid in Agrobacterium tumefaciens essential for crown gall-inducing ability. Nature 252, 169–170.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2006 Humana Press Inc., Totowa, NJ
About this protocol
Cite this protocol
Dandekar, A.M., Teo, G., Uratsu, S.L., Tricoli, D. (2006). Apple (Malus × domestica). In: Wang, K. (eds) Agrobacterium Protocols Volume 2. Methods in Molecular Biology, vol 344. Humana Press. https://doi.org/10.1385/1-59745-131-2:253
Download citation
DOI: https://doi.org/10.1385/1-59745-131-2:253
Publisher Name: Humana Press
Print ISBN: 978-1-58829-843-0
Online ISBN: 978-1-59745-131-4
eBook Packages: Springer Protocols