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Grafting as a Research Tool

  • Colin G.N. Turnbull
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 655)

Abstract

Grafting as a means to connect different plant tissues has been enormously useful in many studies of long-distance signalling and transport in relation to regulation of development and physiology. There is an almost infinite number of pairwise graft combinations that can be tested, typically between two different genotypes and/or between plants previously exposed to different environmental treatments. Grafting experiments are especially powerful for unambiguous demonstration of spatial separation of source and target, including genetic complementation of mutant phenotypes across a graft union, direct detection of transmitted molecules in receiving tissue or vascular sap, and activation or suppression of molecular targets due to signal transmission. Although grafting has a long history in research, only in the past decade has it been applied extensively to the Arabidopsis model. This chapter compares the main Arabidopsis grafting methods now available and describes seedling grafting in detail. Information is also provided on grafting of other common research model species, together with outlines of some successful applications.

Key words

Grafting long-distance signalling 

Notes

Acknowledgements

I am grateful to Jon Booker and Ottoline Leyser for very substantial contributions to development and refinement of Arabidopsis grafting methods and to Christine Beveridge for expert instruction in pea grafting. Financial support from the The Royal Society and the Gatsby Charitable Foundation enabled the original development of several of the techniques described here.

References

  1. 1.
    Walker, R. R., Blackmore, D. H., Clingeleffer, P. R., and Iacono, F. (1997) Effect of salinity and Ramsey rootstock on ion concentrations and carbon dioxide assimilation in leaves of drip-irrigated, field-grown grapevines (Vitis vinifera L. cv. Sultana). Aust J Grape Wine Res 3, 66–74.CrossRefGoogle Scholar
  2. 2.
    Hartmann, T. H., Kester, E. D., Davies, T. F., and Geneve, L. R. (1997). Plant Propagation: Principles and Practices. Prentice Hall, Englewood Cliffs, NJ.Google Scholar
  3. 3.
    Tournier, B., Tabler, M., and Kalantidis, K. (2006) Phloem flow strongly influences the systemic spread of silencing in GFP Nicotiana benthamiana plants. Plant J 47, 383–394.PubMedCrossRefGoogle Scholar
  4. 4.
    Flaishman, M. A., Loginovsky, K., Golobowich, S., and Lev-Yadun S. (2008) Arabidopsis thaliana as a model system for graft union development in homografts and heterografts. J Plant Growth Regul 27, 231–239.CrossRefGoogle Scholar
  5. 5.
    Roney, J. K., Khatibi, P. A., and Westwood, J. H. (2007) Cross-species translocation of mRNA from host plants into the parasitic plant dodder. Plant Physiol 143, 1037–1043.PubMedCrossRefGoogle Scholar
  6. 6.
    David-Schwartz, R., Runo, S., Townsley, B., Machuka, J., and Sinha, N. (2008). Long-distance transport of mRNA via parenchyma cells and phloem across the host-parasite junction in Cuscuta. New Phytol 179, 1133–1141.PubMedCrossRefGoogle Scholar
  7. 7.
    Kaddoura, R. L. and Mantell, S. H. (1991) Synthesis and characterization of Nicotiana-Solanum graft chimeras. Ann Bot 68, 547–556.Google Scholar
  8. 8.
    Tiedemann, R. (1989). Graft union development and symplastic phloem contact in the heterograft Cucumis sativus on Cucurbita ficifolia. J Plant Physiol 134, 427–440.CrossRefGoogle Scholar
  9. 9.
    Ruiz-Medrano, R., Xoconostle-Cazares, B., and Lucas, W. J. (1999) Phloem long-distance transport of CmNACP mRNA: Implications for supracellular regulation in plants. Development 126, 4405–4419.PubMedGoogle Scholar
  10. 10.
    Davis, A. R., Perkins-Veazie, P., Sakata, Y., López-Galarza, S., Maroto, J. V., Lee, S. G, Huh, Y. C., Sun, Z., Miguel, A., King, S. R., Cohen, R., and Lee, J. M. (2008). Cucurbit Grafting. Crit Rev Plant Sci 27, 50–74.CrossRefGoogle Scholar
  11. 11.
    Kubota, C., McClure, M. A., Kokalis-Burelle, N., Bausher,M. G., and Rosskopf, E. N. (2008) Vegetable grafting: History, use and current technology status in North America. HortScience 43, 1664–1669.Google Scholar
  12. 12.
    Delves, A. C., Mathews, A., Day, D. A., Carter, A. S., Carroll, B. J., and Gresshoff, P. M. (1985) Regulation of the soybean–rhizobium nodule symbiosis by shoot and root factors. Plant Physiol 82, 588–590.CrossRefGoogle Scholar
  13. 13.
    Searle, I. R., Men, A. E., Laniya, T. S., Buzas, D. M., Iturbe-Ormaetxe, I., Carroll, B. J., and Gresshoff, P. M. (2003) Long-distance signaling in nodulation directed by a CLAVATA1-like receptor kinase. Science 299, 109–112.PubMedCrossRefGoogle Scholar
  14. 14.
    Oka-Kira, E. and Kawaguchi, M. (2006) Long-distance signaling to control root nodule number. Curr Opin Plant Biol 9, 496–502.PubMedCrossRefGoogle Scholar
  15. 15.
    Foo, E., Turnbull, C. G. N., and Beveridge, C. A. (2001) Long-distance signaling and the control of branching in the rms1 mutant of pea. Plant Physiol 126, 203–209.PubMedCrossRefGoogle Scholar
  16. 16.
    Turnbull, C. G. N., Booker, J. P., and Leyser, H. M. O. (2002) Micrografting techniques for testing long-distance signalling in Arabidopsis. Plant J 32, 255–262.PubMedCrossRefGoogle Scholar
  17. 17.
    Rhee, S. Y. and Somerville, C. R. (1995) Flat-surface grafting in Arabidopsis thaliana. Plant Mol Biol Rep. 13, 118–123.CrossRefGoogle Scholar
  18. 18.
    Ayre, B. G. and Turgeon, R. (2004) Graft transmission of a floral stimulant derived from CONSTANS. Plant Physiol 135, 2271–2278.PubMedCrossRefGoogle Scholar
  19. 19.
    Chen A., Komives, E. A., and Schroeder, J. I. (2006) An improved grafting technique for mature Arabidopsis plants demonstrates long-distance shoot-to-root transport of phytochelatins in Arabidopsis. Plant Physiol 141, 108–120.PubMedCrossRefGoogle Scholar
  20. 20.
    Bainbridge, K., Bennett, T., Turnbull, C., and Leyser, O. (2006) Grafting. In: Arabidopsis Protocols, 2nd edition, Methods in Molecular Biology Volume 323, pp. 39–44. Salinas, J. and Sanchez-Serrano, J.J., eds., Humana Press, Totowa, NJ, ISBN: 1-58829-395-5.Google Scholar
  21. 21.
    An, H.L., Roussot, C., Suarez-Lopez, P., Corbesier, L., Vincent, C., Pineiro, M., Hepworth, S., Mouradov, A., Justin, S., Turnbull, C., and Coupland, G. (2004) CONSTANS acts in the phloem to regulate a systemic signal that induces photoperiodic flowering of Arabidopsis. Development 131, 3615–3626.PubMedCrossRefGoogle Scholar
  22. 22.
    Corbesier, L., Vincent, C., Jang, S., Fornara, F., Fan, Q., Searle, I., Giakountis, A., Farrona, S., Gissot, L., Turnbull, C., and Coupland, G. (2007) FT protein movement contributes to long-distance signaling in floral induction of Arabidopsis. Science 316, 1030–1033.PubMedCrossRefGoogle Scholar
  23. 23.
    Notaguchi, M., Abe, M., Kimura, T., Daimon, Y., Kobayashi,T., Yamaguchi, A., Tomita, Y., Dohi, K., Mori, M., and Araki, T. (2008) Long-distance, graft-transmissible action of Arabidopsis FLOWERING LOCUS T protein to promote flowering. Plant Cell Physiol 49, 1645–1658.PubMedCrossRefGoogle Scholar
  24. 24.
    Green, L. S. and Rogers, E. E. (2004) FRD3 controls iron localization in Arabidopsis. Plant Physiol 136, 2523–2531.PubMedCrossRefGoogle Scholar
  25. 24.
    Van Norman, J. M., Frederick, R. L., and Sieburth, L. E. (2004) BYPASS1 negatively regulates a root-derived signal that controls plant architecture. Curr Biol 14, 1739–1746.PubMedCrossRefGoogle Scholar
  26. 26.
    Rus, A., Baxter, I., Muthukumar, B., Gustin, J., Lahner, B., Yakubova, E., and Salt, D. E. (2006) Natural variants of AtHKT1 enhance Na+ accumulation in two wild populations of Arabidopsis. PLOS Genet 2, 1964–1973.CrossRefGoogle Scholar
  27. 27.
    Bari, R., Pant, B. D., Stitt, M., and Scheible, W. R. (2006) PHO2, microRNA399, and PHR1 define a phosphate-signaling pathway in plants. Plant Physiol 141, 988–999.PubMedCrossRefGoogle Scholar
  28. 28.
    Xia, Y. J., Suzuki, H., Borevitz, J., Blount, J., Guo, Z. J., Patel, K., Dixon, R. A., and Lamb, C. (2004) An extracellular aspartic protease functions in Arabidopsis disease resistance signaling. EMBO J 23, 980–988.PubMedCrossRefGoogle Scholar
  29. 29.
    Brosnan, C. A., Mitter, N., Christie, M., Smith, N. A., Waterhouse, P. M., and Carroll, B. J. (2007) Nuclear gene silencing directs reception of long-distance mRNA silencing in Arabidopsis. Proc Natl Acad Sci USA 104, 14741–14746.PubMedCrossRefGoogle Scholar
  30. 30.
    Pant, B. D., Buhtz, A., Kehr, J., and Scheible, W .R. (2008) MicroRNA399 is a long-distance signal for the regulation of plant phosphate homeostasis. Plant J 53, 731–738.PubMedCrossRefGoogle Scholar
  31. 31.
    Wilmoth, J. C., Wang, S. C., Tiwari, S. B., Joshi, A. D., Hagen, G., Guilfoyle, T. J., Alonso, J. M., Ecker, J. R., and Reed, J. W. (2005) NPH4/ARF7 and ARF19 promote leaf expansion and auxin-induced lateral root formation. Plant J 43, 118–130.PubMedCrossRefGoogle Scholar
  32. 32.
    Foo, E., Morris, S. E., Parmenter, K., Young, N., Wang, H., Jones, A., Rameau, C., Turnbull, C. G. N., and Beveridge, C.A. (2007) Feedback Regulation of xylem cytokinin content is conserved in pea and Arabidopsis. Plant Physiol 143, 1418–1428.PubMedCrossRefGoogle Scholar
  33. 33.
    Matsumoto-Kitano, M., Kusumoto, T., Tarkowski,P., Kinoshita-Tsujimura, K., Vaclavikova, K., Miyawaki, K., and Kakimoto, T. (2008) Cytokinins are central regulators of cambial activity. Proc Natl Acad Sci USA 105, 20027–20031.PubMedCrossRefGoogle Scholar
  34. 34.
    Christmann, A., Weiler, E. W., Steudle, E., and Grill, E. (2007) A hydraulic signal in root-to-shoot signalling of water shortage. Plant J 52, 167–174.PubMedCrossRefGoogle Scholar
  35. 35.
    Wilson, A. K., Pickett, F. B., Turner, J. C., and Estelle, M. (1990) A dominant mutation in Arabidopsis confers resistance to auxin, ethylene and abscisic acid. Mol Gen Genet 222, 377–383.PubMedCrossRefGoogle Scholar
  36. 36.
    Napoli, C. (1996) Highly branched phenotype of the petunia dad1-1 mutant is reversed by grafting. Plant Physiol 111, 27–37.PubMedGoogle Scholar
  37. 37.
    Murfet, I. C. (1971) Flowering in Pisum: Reciprocal grafts between known genotypes. Aust J Biol Sci 24, 1089–1101.Google Scholar
  38. 38.
    Beveridge, C. A., Ross, J. J., and Murfet, I. C. (1994) Branching mutant rms-2 in Pisum sativum – grafting studies and endogenous indole-3-acetic-acid levels. Plant Physiol 104, 953–959.PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Colin G.N. Turnbull
    • 1
  1. 1.Division of BiologyImperial College LondonLondonUK

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