Molecular Breeding

, Volume 4, Issue 2, pp 99–109 | Cite as

Molecular analysis of the genome of transgenic rice (Oryza sativa L.) plants produced via particle bombardment or intact cell electroporation

  • Ariel Arencibia
  • Eugenio Gentinetta
  • Elena Cuzzoni
  • Stefano Castiglione
  • Aiay Kohli
  • Philippe Vain
  • Mark Leech
  • Paul Christou
  • Francesco SalaEmail author


In the present work we utilised some of the most discriminative molecular tools, such as RAPD, AFLP, AFRP and RAMP, to analyse the genome of independently derived transgenic plants from three elite Italian cultivars (cv. Lido, Carnaroli and Thaibonnet) and found that two methods for direct gene transfer, namely particle bombardment and intact cell electroporation (the latter being a procedure set up in this work), result in transgenic rice (Oryza sativa L.) plants that exhibit negligible genomic changes. This is in contrast with recently published results showing relevant changes in the DNA of transgenic rice plants generated through protoplasts electroporation and of transgenic poplar plants engineered through Agrobacterium tumefaciens infection. Implications of these findings are discussed in the context of selecting appropriate gene transfer methodologies to produce transgenic plants expressing genes of interest while retaining their genomic integrity and, thus, their superior agronomic and/or industrial traits.

transgenic rice particle bombardment cell electroporation RAPD AFLP AFRP RAMP 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    Arencibia A, Molina P, De la Riva G, Selman-Housein G: Production of transgenic sugarcane (Saccharum officinarum L.) plants by intact cell electroporation. Plant Cell Rep 14: 305–309 (1995).Google Scholar
  2. 2.
    Bao PH, Granata S, Castiglione S, Wang G, Giordani C, Cuzzoni E, Damiani G, Bandi C, Datta SK, Datta K, Potrykus I, Callegarin A, Sala F: Evidence for genomic changes in transgenic rice (Oryza sativa L.) recovered from protoplasts. Transgen Res 5: 97–103 (1996).Google Scholar
  3. 3.
    Bellini C, Guerche A, Spielman J, Goujaud C, Lesaint C, Caboche M: Genetic analysis of transgenic tobacco plants obtained by liposo-memediated transformation: absence of evidence for themutagenic effect of inserted sequences in sixty characterised transformants. J Hered 80: 361–367 (1989).Google Scholar
  4. 4.
    Bellini C, Giordani C, Lupotto E, Locatelli F, Cuzzoni E, Avogadro E, Castiglione S, Sala F: Stability of a foreign gene in transgenic Nicotiana tabacum L. plants during a cycle of dedifferentiation/ differentiation. Plant Sci 82: 193–200 (1992).Google Scholar
  5. 5.
    Castiglione S, Wang G, Damiani G, Bandi C, Bisoffi S, Sala F: RAPD fingerprints for the identification of elite poplar (Populus spp.) clones. Theor Appl Genet 85: 54–59 (1993).Google Scholar
  6. 6.
    Chalhoub BA, Thibault S, Laucou V, Rameau C, Cousin R: Silver staining and recovery of AFLP amplification products on large denaturing polyacrylamide gels. BioTechniques 22: 216–220 (1997).Google Scholar
  7. 7.
    Chan MT, Chang HH, Ho SL, Tong WF, Yu SM: Agrobacterium-mediated production of transgenic rice plants expressing a chimeric α-amylase promoter β-glucuronidase gene. Plant Mol Biol 22: 491–506 (1993).Google Scholar
  8. 8.
    Christou P, Swain WF, Yang NS, McCabe DE: Inheritance and expression of foreign genes in transgenic soybean plants. Proc Natl Acad Sci USA 86: 7500–7504 (1989).Google Scholar
  9. 9.
    Christou P, Ford TL, Kofron M: Production of transgenic rice (Oryza sativa L.) plants from agronomically important Indica and Japonica varieties via electric discharge particle acceleration of exogenous DNA into immature zygotic embryos. Bio/technology 9: 957–962 (1991).Google Scholar
  10. 10.
    Christou P: Genetic transformation of crop plants using microprojectile bombardment. Plant J 2: 275–281 (1992).Google Scholar
  11. 11.
    Christou P, Ford TL and Kofron M: The development of a varietyin-dependent gene-transfer method for rice. Trends Biotechnol 10: 239–246 (1992).Google Scholar
  12. 12.
    Cooley J, Ford T, Christou P: Molecular and genetic characterisation of elite transgenic rice plants produced by electricdischarge particle acceleration. Theor Appl Genet 90: 97–104 (1995).Google Scholar
  13. 13.
    Dale PJ, McPartland HC: Field performance of transgenic potato plants compared with controls regenerated from tuber discs and shoot cuttings. Theor Appl Genet 84: 585–591 (1992).Google Scholar
  14. 14.
    Dale PJ, Irwin JA, Scheffler JA: The experimental and commercial release of transgenic crop plants. Plant Breed 3: 1–22 (1993).Google Scholar
  15. 15.
    Datta SK, Peterhans A, Datta K, Potrykus I: Genetically engineered fertile indica-rice recovered from protoplasts. Bio/technology 8: 736–740 (1990).Google Scholar
  16. 16.
    De Block M, Butterman J, Wandewile M, Dockx J, Thoen C, Gossele V, Rao Movva N, Thompson C, Van Montague M, Leemans J: Engineering herbicide resistance in plants by expression of a detoxifying enzyme. EMBO J 6: 2513–2518 (1987).Google Scholar
  17. 17.
    Dekeyser RA, Claes B, De Rycke RMU, Habets ME, Van Montague MC, Caplan AB: Transient gene expression in intact and organized rice tissue. Plant Cell 2: 592–600 (1990).Google Scholar
  18. 18.
    Gheysen G, Dhaese P, Van Montague M, Schell J: DNA flux across genetic barriers: the crown gall phenomenon. In: Hohn B, Dennis E (eds) Plant Gene Research, Genetic flux in Plants, pp. 12–47. Springer-Verlag, Vienna, (1985).Google Scholar
  19. 19.
    Hiei Y, Ohta S, Komari T, Kumashiro T: Efficient transformation of rice (Oryza sativa L.) mediated by Agrobacterium and sequence analysis of the boundaries of the TDNA. Plant J 6: 271–282 (1994).Google Scholar
  20. 20.
    Jefferson RA, Kavanagh TA, Bevan MW: GUS fusion: β-glucuronidase as a sensitive and versatile gene fusion marker in plants. EMBO J 6: 3901–3907 (1987).Google Scholar
  21. 21.
    Karp A: On the current understanding of somaclonal variation. Oxford Surv Plant Mol Cell Biol 7: 1–58 (1991).Google Scholar
  22. 22.
    Karp A, Ingram DS, Isaac PG (eds) MolecularTools for Screening Biodiversity: Plants and Animals. Chapman and Hall, London (1998).Google Scholar
  23. 23.
    Larkin PJ, Scowcroft WR: Somaclonal variation. A novel source of variability from cell cultures for plant improvement. Theor Appl Genet 60: 197–214 (1981).Google Scholar
  24. 24.
    Lee N, Wang Y, Yang J, Ge K, Huang S, Tan, Testa D: Efficient transformation and regeneration of rice small cell groups. Proc Natl Acad Sci USA 88: 6389–6393 (1991).Google Scholar
  25. 25.
    Murashige T, Skoog F: A revised medium for rapid growth and bioassay with tobacco tissue culture. Physiol Plant 15: 473–497 (1962).Google Scholar
  26. 26.
    Paszkowski J, Shillito RD, Saul M, Mandak V, Hohn T, Hohn B, Potrykus I: Direct gene transfer to plants. EMBO J 3: 2717–2722 (1984).Google Scholar
  27. 27.
    Rhodes CA, Pierce DA, Mettler IJ, Mascarenhas D, Detmer JJ: Genetically transformed maize plants from protoplasts. Science 240: 204–207 (1988).Google Scholar
  28. 28.
    Schuh W, Nelson MR, Bigelow DM, Orum TV, Orthj CE, Lynch PT, Eyles PS, Blackhall NW, Jones J, Cocking EC, Davey MR: The phenotypic characterisation of R2 generation transgenic rice plants under field conditions. Plant Sci 89: 69–79 (1993).Google Scholar
  29. 29.
    Tauts D: Hypervariability of simple sequences as a general source for polymorphic DNA markers. Nucleic Acids Res 17: 6463–6471 (1989).Google Scholar
  30. 30.
    Vos P, Hogers R, Bleeker M, Reijans M, van de Lee T, Hornes M, Frijters A, Pot J, Peleman J, Kuiper M, Zabeau M: AFLP: a new technique for DNA fingerprinting. Nucl Acids Res 23: 4407–4414 (1995).Google Scholar
  31. 31.
    Wang G, Castiglione S, Zhang J, Fu R, Ma J, Li W, Sun Y, Sala F: Hybrid rice (Oryza sativa L.): identification and parentage determination by RAPD fingerprinting. Plant Cell Rep 14: 112–115 (1994).Google Scholar
  32. 32.
    Wang G, Castiglione S, Chen Y, Li L, Han Y, Tian Y, Gabriel DW, Han Y, Mang K, Sala F.: Poplar (Populus nigra L.) Plants transformed with a Bacillus thuringiensis toxin gene: insecticidal activity and genome analysis. Transgen Res 5: 289–301 (1996).Google Scholar
  33. 33.
    Wu K, Jones R, Danneberger L, Scolnik P: Detection of microsatellite polymorphisms without cloning. Nucl Acids Res 22: 3257–3258 (1994).Google Scholar
  34. 34.
    Xu Y, Bu W, Li B: Metabolic factors capable of inducing Agrobacterium vir gene expression are present in rice (Oryza sativa L.). Plant Cell Rep 12: 160–164 (1993).Google Scholar
  35. 35.
    Zhang HM, Yang H, Rech EL, Golds TJ, Davis AS, Mulligan BJ, Cocking EC, Davey MR: Transgenic rice plants produced by electroporation-mediated plasmid uptake into protoplasts. Plant Cell Rep 7: 379–384 (1988).Google Scholar

Copyright information

© Kluwer Academic Publishers 1998

Authors and Affiliations

  • Ariel Arencibia
    • 1
  • Eugenio Gentinetta
    • 2
  • Elena Cuzzoni
    • 1
  • Stefano Castiglione
    • 1
  • Aiay Kohli
    • 3
  • Philippe Vain
    • 3
  • Mark Leech
    • 3
  • Paul Christou
    • 3
  • Francesco Sala
    • 1
    Email author
  1. 1.Dipartimento di BiologiaUniversità di Milano20133Italia
  2. 2.Centro Ricerche sul Riso. Castello d'AgognaPaviaItalia
  3. 3.John Innes Centre. Norwich Research ParkColney, NorwichUK

Personalised recommendations