Regeneration and Transformation Experiments in Apple

  • Margareta Welander
  • Gowri Maheswaran
Part of the NATO ASI Series book series (NSSA, volume 210)


Successful introduction of foreign genes using a leaf disc procedure and cocultivation with Aqrobacterium tumefaciens requires a good regeneration protocol and tissue susceptible to infection. In our experiments, apple rootstocks M26 and M9 and the cultivar Mcintosh Wijcik were used. A good regeneration system was developed for M9 whereas a suitable protocol for the others had been previously developed. Transformation experiments were carried out with three wild type strains and three disarmed vectors harbouring ß-glucuronidase and neomycin phosphotransferase genes. Infection with three wild type strains (C58, A348, A281) did not cause gall formation. However, the binary vector pBH21 (35SGUS) x LBA 4404 resulted in transformed shoots of M26 and transformed callus of M9. This was confirmed histochemically using the substrate 5-bromo-4-chloro-3-indolyl glucoronide (X-Gluc) staining. Cefotaxime, used to control growth of the bacteria, had an effect simalar to cyto-kinin by stimulating shoot formation in vitro.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    Alston, F.H., 1970, Resistance to collar rot. Phytophthora cactorum (Leb.and Cohn) Schroet.,in apple. Rep. E.Mailing Res. Stn. for 1969: 143–145.Google Scholar
  2. 2.
    Byrne, M.C., McDonnel, R.E., Wright, M.S., and Carnes, M.G., 1987, Strain and cultivar specificity in the Aqrobacterium-soybean interaction. Plant Cell Tiss. Org. Cult. 8: 3–15.CrossRefGoogle Scholar
  3. 3.
    Chu, C.C., Wang, C.C., Sun, C.S., Hsü, C., Yin, K.C., Chu, C.Y., and Bi, F.Y., 1975, Establishment of an efficient medium for anther culture of rice through comparative experiments on the nitrogen sources. Sci. Sin. 18: 659–668.Google Scholar
  4. 4.
    Debergh, P., Harbaoui, Y., and Lemeur, R., 1981, Mass propagation of globe artichoke (Cynara scolymus): Evaluation of different hypothesis to overcome vitrification with special reference to water potential. Physiol. Plant. 53: 181–187.CrossRefGoogle Scholar
  5. 5.
    Gamborg, O.L., Miller, R.A., and Ojima, K., 1968, Nutrient requirements of suspension cultures of soybean root cells. Exp. Cell Res. 50: 151–158.CrossRefPubMedGoogle Scholar
  6. 6.
    Hooykaas-Van Slogteren, G.M.S., Hooykaas, P.J.J., and Schilperoort, R.A., 1984, Expression of Ti plasmid genes in monocotyledonous plants infected with Aqrobacterium tumefaciens. Nature vol 311 (5988): 763–764.CrossRefGoogle Scholar
  7. 7.
    James, D.J., 1987, Cell and tissue culture technology for the genetic manipulation of temperate fruit trees. Biotechnology and Genetic Reviews vol.5: 33–79.CrossRefGoogle Scholar
  8. 8.
    James, D.J., Passey, A.J., Barbara, D.J., and Bevan, M., 1989, Genetic transformation of apple (Malus pumila Mill) using a disarmed Ti-binary vector. Plant Cell Reports 7: 658–661.PubMedGoogle Scholar
  9. 9.
    Jefferson, R.A., Burgess, S.M., and Hirsh, D., 1986, ß-Glucuronidase from E.coli as a gen fusion marker. Proc. Natl. Acad. Sci.USA 83: 8447–51.CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Klee, H., Horsch, R., and Rogers, S., 1987, Aqrobacterium-mediated plant transformation and its further applications to plant biology. Ann. Rev. Plant. Physiol. 38: 467–486.CrossRefGoogle Scholar
  11. 11.
    Knight, R.L., and Alston, F.H., 1969, Developments in apple breeding. Rep. E.Mailing Res. Stn. for 1968: 125–132.Google Scholar
  12. 12.
    Mathews, H., 1988, In vitro responses of Brassica juncea and Viqna radiata to the antibiotic kanamycin. Ann. Bot. 62: 671–675.CrossRefGoogle Scholar
  13. 13.
    Mathias, R.J., and Boyd, L.,1986, Cefotaxime stimulates callus growth, embryogenesis and regeneration in hexaploid bread wheat (Triticum aestivum L EM. Thell). Plant Sci. 46: 217–223.CrossRefGoogle Scholar
  14. 14.
    Mathias, R.J., and Mukasa, C., 1987, The effect of cefotaxime on the growth and regeneration of callus from four varieties of barley (Hordeum vulgare L.). Plant Cell Reports 6: 454–457.PubMedGoogle Scholar
  15. 15.
    Murashige, T., and Skoog, F., 1962, A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol. Plant. 15: 473–497.CrossRefGoogle Scholar
  16. 16.
    Pasqualetto, P.L., Zimmerman R.H., and Fordham, I., 1986, Gelling agent and growth regulator effects on shoot vitrification of “Gala” apple in vitro. J. Amer. Soc. Hort. Sci. 111(6): 976–980.Google Scholar
  17. 17.
    Pierik, R.L.M., 1971, Plant tissue culture as motivation for the symposium. Misc. Pap. Landbouwhogesch. Wageningen. 9: 3–13.Google Scholar
  18. 18.
    Romberger, J.A., and Tabor, C.A., 1971, The Picea abies shoot apical meristem in culture. I. Agar and autoclaving effects. Amer. J. Bot. 58(2): 131–140.CrossRefGoogle Scholar
  19. 19.
    Shay, J.R., Dayton, D.F., and Hough, L.F., 1953, Apple scab resistance from a number of Malus species. Proc. Soc. Hort. Sci. 62: 348–356.Google Scholar
  20. 20.
    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.CrossRefGoogle Scholar

Copyright information

© Plenum Press, New York 1991

Authors and Affiliations

  • Margareta Welander
    • 1
  • Gowri Maheswaran
    • 2
  1. 1.Department of Horticultural ScienceAlnarpSweden
  2. 2.Horticultural Research InstituteKnoxfield, Ferntree GullyAustralia

Personalised recommendations