Agroinoculation: A Simple Procedure for Systemic Infection of Plants with Viruses

  • Zarir E. Vaghchhipawala
  • Kirankumar S. Mysore
Part of the Methods in Molecular Biology™ book series (MIMB, volume 451)


Plant-virus interaction studies, for long, plagued by asynchronous/failed infections, have improved since the usage of Agrobacterium as a delivery agent for viral genomes. Popularly known as “agroinoculation,” this method has revolutionized plant virology studies, leading to identification of viruses as casual agents of disease, viral genome mutagenesis and recombination analyses, and virus-induced gene silencing (VIGS) applications. We present here a brief overview of the recent applications of this method and a detailed protocol for agroinoculation and VIGS used in our laboratory.


Agrobacterium Agroinoculation Agroinfiltration Agrodrench VIGS Tobacco rattle virus (TRV) vector 


  1. 1.
    1. Grimsley, N., Hohn, B., Hohn, T., and Walden, R. (1986) “Agroinfection,“ an alternative route for viral infection of plants by using the Ti plasmid. Proc Natl Acad Sci 83, 3282–86.PubMedCrossRefGoogle Scholar
  2. 2.
    2. Grimsley, N., Hohn, T., Davies, J. W., and Hohn, B. (1987) Agrobacterium-mediated delivery of infectious maize streak virus into maize plants. Nature 325, 177–79.CrossRefGoogle Scholar
  3. 3.
    3. Elmer, J. S., Sunter, G., Gardiner, W. E., Brand, L., Browning, C. K., Bisaro, D. M., and Rogers, S. G. (1988) Agrobacterium-mediated inoculation of plants with tomato golden mosaic virus DNAs. Plant Mol Biol 10, 225–34.CrossRefGoogle Scholar
  4. 4.
    4. Navot, N., Pichersky, E., Zeidan, M., Zamir, D., and Czosnek, H. (1991) Tomato yellow leaf curl virus: A whitefly-transmitted geminivirus with a single genomic component. Virology 185, 151–61.PubMedCrossRefGoogle Scholar
  5. 5.
    5. Czosnek, H., Kheyr-Pour, A., Gronenbom, B., Remetz, E., Zeidan, M., Altman, A., Rabinowitch, H. D., Vidavsky, S., Kedar, N., Gafni, Y., and Zamir, D. (1993) Replication of tomato yellow leaf curl virus (TYLCV) DNA in agroinoculated leaf discs from selected tomato genotypes. Plant Mol Biol 22, 995–1005.PubMedCrossRefGoogle Scholar
  6. 6.
    6. Bendahmane, M., Schalk, H.-J., and Gronenborn, B. (1995) Identification and characterization of wheat dwarf virus from France using a rapid method for geminiviurs DNA preparation. Mol Plant Pathol 85, 1449–55.Google Scholar
  7. 7.
    7. Buragohain, A., Sung, Y., Coffin, R., and Coutts, R. (1994) The infectivity of dimeric potato yellow mosaic geminivirus clones in different hosts. J Gen Virol 75, 2857–61.PubMedCrossRefGoogle Scholar
  8. 8.
    8. Mandal, B., Varma, A., and Malathi, V. G. (1997) Systemic infection of Vigna mungo using the cloned DNAs of the blackgram isolate of mungbean yellow mosaic geminivirus through agroinoculation and transmission of the progeny virus by whiteflies. J Phytopathol 145, 505–10.CrossRefGoogle Scholar
  9. 9.
    9. Lamprecht, S., and Jelkmann, W. (1997) Infectious cDNA clone used to identify strawberry mild yellow edge-associated potexvirus as causal agent of the disease. J Gen Virol. 78, 2347–53.PubMedGoogle Scholar
  10. 10.
    10. Klinkenberg, F. A., EUwood, S., and Stanley, J. (1989) Fate of African cassava Mosaic virus coat protein deletion mutants after agroinoculation. J Gen Virol 70, 1837–44.CrossRefGoogle Scholar
  11. 11.
    11. Stanley, J., Latham, J. R., Pinner, M. S., Bedford, 1., and Markham, P G. (1992) Mutational analysis of the monopartite geminivirus beet curly top virus. Virology 191, 396–405.PubMedCrossRefGoogle Scholar
  12. 12.
    12. Sung, Y, and Coutts, R. (1995) Mutational analysis of potato yellow mosaic geminivirus. J Gen Virol 76, 1773–80.PubMedCrossRefGoogle Scholar
  13. 13.
    13. Sadowy, E., Maasen, A., Juszczuk, M., David, C, Zagorski-Ostoja, W., Gronenborn, B., and Hulanicka, M. D. (2001) The OREO product of Potato leafroU virus is indispensable for virus accumulation. J Gen Virol 82, 1529–32.PubMedGoogle Scholar
  14. 14.
    14. Sadowy, E., Juszczuk, M., David, C, Gronenborn, B., and Hulanicka, M. D. (2001) Mutational analysis of the proteinase function of Potato leafroU virus. J Gen Virol 82, 1517–27.PubMedGoogle Scholar
  15. 15.
    15. Boulton, M. I., King, D. I., Markham, P G., Pinner, M. S., and Davies, J. W. (1991) Host range and symptoms are determined by specific domains of the maize streak virus genome. Virology 181,312–18.PubMedCrossRefGoogle Scholar
  16. 16.
    16. Rigden, J. E., Krake, L. R., Rezaian, M. A., and Dry, I. B. (1994) ORE C4 of tomato leaf curl geminivirus is a determinant of symptom severity. Virology 204, 847–50.PubMedCrossRefGoogle Scholar
  17. 17.
    17. Saunders, K., and Stanley, J. (1995) Complementation of African cassava mosaic virus AC2 gene function in a mixed bipartite geminivirus infection. J Gen Virol 76, 2287–92.PubMedCrossRefGoogle Scholar
  18. 18.
    18. Sung, Y, and Coutts, R. (1995) Pseudorecombination and complementation between potato yellow mosaic geminivirus and tomato golden mosaic geminivirus. J Gen Virol 76, 2809–15.PubMedCrossRefGoogle Scholar
  19. 19.
    19. Kiraly, L., Bourque, J. E., and Schoelz, J. E. (1998) Temporal and spatial appearance of recombinant viruses formed between cauliflower mosaic virus (CaMV) and CaMV sequences present in transgenic Nicotiana bigelovii. Mol Plant Microbe Interact 11, 309–16.CrossRefGoogle Scholar
  20. 20.
    20. Kheyr-Pour, A., Gronenborn, B., and Czosnek, H. (1994) Agroinoculation of tomato yellow leaf curl vims (TYLCV) overcomes the virus resistance of wild Lycopersicon species. Plant Breed 112, 228–33.CrossRefGoogle Scholar
  21. 21.
    21. Cruz, F. C. S., Boulton, M. 1., Hull, R., and Azzam, O. (1999) International Rice Research Institute, Los Banos, Philippines. J Phytopathol 147, 653–59.CrossRefGoogle Scholar
  22. 22.
    22. Martin, B. P., Willment, J. A., and Rybicki, E. P. (1999) Evaluation of maize streak virus pathogenicity in differentially resistant Zea mays genotypes. Virology 89, 695–700.Google Scholar
  23. 23.
    23. Martin, B. P., and Rybicki, E. P. (2000) Improved efficiency of Zea mays agroinoculation with Maize streak virus. Plant Dis 84, 1096–98.CrossRefGoogle Scholar
  24. 24.
    24. Tripathi, S., and Varma, A. (2003) Identification of sources of resistance in Lycopersicon species to Tomato leaf curl geminivirus (ToLCV) by agroinoculation. Euphytica 129, 43–52.CrossRefGoogle Scholar
  25. 25.
    25. Ding, X. S., Liu, J., Chen, N.-H., Fohmonov, A., Hou, Y.-M., Bao, Y, Katagi, C, Carter, S. A., and Nelson, R. S. (2004) The Tobacco mosaic virus 126-kDa protein associated with virus replication and movement suppresses RNA silencing. Mol Plant Microbe Interact 17, 583–92.PubMedCrossRefGoogle Scholar
  26. 26.
    26. Kościanśka, E., Kalantidis, K., Wypijewski, K., Sadowski, J., and Tabler, M. (2005) Analysis of RNA Silencing in Agroinfiltrated Leaves of Nicotiana benthamiana and Nicotiana tabacum. Plant Mol Biol 59, 647–61.PubMedCrossRefGoogle Scholar
  27. 27.
    27. Ryabov, E. V., van Wezel, R., Walsh, J., and Hong, Y (2004) Cell-to-cell, but not long-distance, spread of RNA silencing that is induced in individual epidermal cells. J Virol 78, 3149–54.PubMedCrossRefGoogle Scholar
  28. 28.
    28. Xie, Q., and Guo, H.-S. (2006) Systemic antiviral silencing in plants. Virus Res 118, 1–6.PubMedCrossRefGoogle Scholar
  29. 29.
    29. Herr, A. J., and Baulcombe, D. C. (2004) RNA silencing pathways in plants. Cold Spring Harb Symp Quant Biol 69, 363–70.PubMedCrossRefGoogle Scholar
  30. 30.
    30. Liu, Y, Schiff, M., and Dinesh-Kumar, S. P. (2002) Virus-induced gene silencing in tomato. Plant J 31,777–86.PubMedCrossRefGoogle Scholar
  31. 31.
    31. Dinesh-Kumar, S. P, Anandalakshmi, R., Marathe, R., Schiff, M., and Liu, Y (2003) in “Plant Functional Genomics” (Grotewold, E., Ed.), Vol. 236, pp. 287–93, Humana Press, Inc., Totowa, NJ.CrossRefGoogle Scholar
  32. 32.
    32. Ryu, C.-M., Anand, A., Kang, L., and Mysore, K. S. (2004) Agrodrench: A novel and effective agroinoculation method for virus-induced gene silencing in roots and diverse Solanaceous species. Plant J 40, 322–31.PubMedCrossRefGoogle Scholar

Copyright information

© Humana Press, a part of Springer Science + Business Media, LLC 2008

Authors and Affiliations

  • Zarir E. Vaghchhipawala
    • 1
  • Kirankumar S. Mysore
    • 1
  1. 1.Plant Biology DivisionThe Samuel Roberts Noble FoundationArdmoreUSA

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