Journal of Forestry Research

, Volume 13, Issue 2, pp 91–97 | Cite as

Genetic transformation ofPinus taeda by particle bombardment

  • Tang Wei
  • Vanessa Samuels


A protocol is presented for genetically engineering loblolly pine (Pinus taeda L.) using particle bombardment. This protocol enabled the routine transformation of loblolly pine plants that were previously difficult to transform. Mature zygotic embryos were used to be bombarded and to generate organogenic callus and transgenic regenerated plants. Plasmid pB48.215 DNA contained a synthetic Bacillus thuringiensis (B.t.) crylAc coding sequence flanked by the double cauliflower mosaic virus (CaMV) 35S promoter and nopaline synthase (Nos) terminator sequences, and the selectable marker gene, neomycin phosphotransferase II (nptil) controlled by the promoter of the nopaline synthase gene was introduced into loblolly pine tissues by particle bombardment. The transformed tissues were proliferated and selected by kanamycin resistance conferred by the introduced NPTIi gene. Shoot regeneration was induced from the kanamycin-resistant callus, and transgenic plantlets were then produced. The presence of the introduced genes in the transgenic loblolly pine plants was confirmed by polymerase chain reactions (PCR) analysis, by Southern blot analysis, and insect feeding assays. The recovered transgenic plants were acclimatized and then established in soil.


Pinus taeda L. Biolistic transformation Bacillus thuringiensis (B.t.) crylAb Insect feeding bioassay 


Q55 S791.255.04 

Document code





Bacillus thuringiensis


cauliflower mosaic virus


2,4-dichlorophenoxyacetic acid


indoie-3-butyric acid


nopaline synthase


neomycin phosphotransferase II gene


polymerase chain reactions


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  1. Alam M.F., Datta, K., Abrigo, E., Vasquez, A., Senadhira, D., Datta, S.K. 1998. Production of transgenic deepwater indica rice plants expressing a syntheticBacillus thuringiesis crylA(b) gene with enhanced resistance to yellow stem borer [J]. Plant Sci., 135: 25–30.CrossRefGoogle Scholar
  2. Cheng, X.Y., Saroana, R., Kaplan, H., Altosaar, I. 1998. Agrobacterium-transformed rice plants expressing synthetic cry1Ab and cry1Ac genes are highly toxic to striped stem borer and yellow stem borer [J]. Proc Natl Acad Sci USA, 95: 2767–2772.PubMedCrossRefGoogle Scholar
  3. Christou, P., Ford, T., Kofron, M. 1991. Production of transgenic rice (Oriza sativa L.) plants from agronomically important indica and japonica varieties via electric discharge particle acceleration of exogenous DNA into immature zygotic embryos [J]. Bio/Technology, 9: 957–962.CrossRefGoogle Scholar
  4. Christou P, Swain, W., Yang N-S, McCabe, D. 1989. Inheritance and expression of foreign genes in transgenic soybean plants [J]. Proc Natl Acad Sci USA, 86: 7500–7504.PubMedCrossRefGoogle Scholar
  5. Datta, K., Vasquez, A., Tu, J., Torrizo, L., Alam, M.F., Oliva, N., Abrigo, E., Khush, G.S., Datta, S.K. 1998. Constitutive and tissue-specific differential expression of crylA(b) gene in transgeic rice plants conferring enhanced resistance to rice insect pest[J]. Theor. Appl. Genet., 97: 20–30.CrossRefGoogle Scholar
  6. Ellis, D.D., McCabe, D.E., McInniss, Ramachandran, R., Russeil, D.R., Wallace, K.M., Martinell, B.J., Roberts, D.R., Raffa, K.F., McCown, B.H. 1993. Stable transformation of Picea glauca by particle acceleration [J]. Bio/Technology, 11: 84–89.CrossRefGoogle Scholar
  7. Fitch, M., Manshardt, R., Gonsalves, D., Slightom, J., Sanford, J. 1990. Stable transformation of papaya via microprojectile bombardment [J]. Plant Cell Rep., 9: 189–194.Google Scholar
  8. Franche, C., Diouf, D., Le, Q., Bogusz, D., N’Diaye, A., Gherbi, H., Gobe, C., Duhoux, E. 1997. Genetic transformation of the actinorhizal tree Allocasuarina vertlcillata by Agrobacterium tumefaciens [J]. Plant J., 11: 897–904.CrossRefGoogle Scholar
  9. Gheysen, G., van Montagu, M., Zambryski, P. 1987. Integration ofAgrobacterium tumefaciens T-DNA involves rearrangements of target plant DNA sequences [J]. Proc Natl Acad Sci USA, 84: 9006–9010.CrossRefGoogle Scholar
  10. Han, K-H, Ma, C., Strauss, S.H. 1997. Matrix attachment regions enhance transgene expression and transformation in poplar [J]. Transgen Res., 6: 415–420.CrossRefGoogle Scholar
  11. Huang, Y., Diner, A.M., Karnosky, D.F. 1991Agrobacterium rhizogenes-mediated genetic transformation and regeneration of a conifer:Larix deciduas [J]. In Vitro Cell Dev Biol., 27: 201–207.CrossRefGoogle Scholar
  12. Humara, J.M., Lopez, M., Ordas, R.J. 1999.Agrobacterium rhizogenes-mediated transformation of Pinus pinea L. cotyledons: an assessment of factors influencing the efficiency of uidA gene transfer [J]. Plant Cell Rep., 19: 51–58.CrossRefGoogle Scholar
  13. Lida, A., Yamashida, T., Yamada, Y., Morikawa, H. 1991. Efficiency of particle-bombardment-mediated transformation is influenced by cell stage in synchronized cultured cells of tobacco. Plant Physiol., 97:1585–1587.CrossRefGoogle Scholar
  14. Klimaszewska, K., Devantier, Y., lachance, D., Lelu, M.A., Charest, P.J. 1997.Larix laricina (tamarack): somatic embryogenesis and genetic transformation [J]. Can J. For. Res., 27: 538–550.CrossRefGoogle Scholar
  15. Levee, V., Lelu, M.A., Jouanin, L., Cornu, D., Pilate, G. 1997. Agrobacterium tumefaciens-mediated transformation of hybrid larch (Larix kaempferi × L. decidua) and transgenic plant regeneration [J]. Plant Cell Rep., 16: 680–685.CrossRefGoogle Scholar
  16. Li, T.Y., Tian, Y.C., Qing, X.F. 1994. Studies on high-efficient insect resistance transgenic tobacco [J]. China Sinica. (B), 24: 276–268.Google Scholar
  17. McCown, B.H., McCabe, D.E., Russell, D.R., Robison, D.J., Barton, K.A., Raffa, K.F. 1991. Stable transformation ofPopulus and incorporation of pest resistance by discharge particle acceleration [J]. Plant Cell Rep., 9: 950–954.CrossRefGoogle Scholar
  18. Perlak, F.J., Fuchs, R.L., Dean, D.A., McPherson, S.L., Fischoff, D.A. 1991. Modification of the coding sequence enhances plant expression of insect control proteins [J]. Proc Natl Acad Sci USA, 88: 3324–3328.PubMedCrossRefGoogle Scholar
  19. Perlak, F.J., Stone, T.b, Muskopf, Y.M., Petersen, L.J., Parker, G.B., McPherson, S.A., Wyman, J., Love, S., Reed, G., Biever D., Fischoff, D.A. 1993. Genetically improved potatoes: protection from damage by Colorado potato beetle [J]. Plant Mol Biol.,22: 313–321.PubMedCrossRefGoogle Scholar
  20. Sambrook, J, Fritsch, E.F., Mamiatis, T. 1989. Molecular Cloning: a laboratory manual (2nd ed) [M]. New York: Cold Spring Harbor.Google Scholar
  21. Schuler, TH., Poppy, G.M., Kerry, B.R., Denholm, L. 1998. Insectresistant transgeic plants [J]. Trends Biotechnol., 16: 168–175.CrossRefGoogle Scholar
  22. Shin, D.I., Podila, G.K., Huang, Y., Karnosky, D.F. 1994. Transgenic larch expressing genes for herbicide and insect resistance [J]. Can. J. For. Res., 24: 2059–2067.CrossRefGoogle Scholar
  23. Stewart, CN. Jr, Adang, M.J., All, J.N., Boerma, H.R., Cardineau, G., Tucker, D., Parrott, W.A. 1996. Genetic transformation, recovery and characterization of fertile soybean transgenic for a syntheticBacillus thuringiesis crylA(c) gene [J]. Plant Physiol 112: 121–129.PubMedCrossRefGoogle Scholar
  24. Tang, W., Ouyang, F., Guo, Z.C. 1998. Plant regeneration through organogenesis from callus induced from mature zygotic embryos of loblolly pine [J]. Plant Cell Rep., 17: 557–560.CrossRefGoogle Scholar
  25. Tzfira, T., Yamitzky, O., Vainstein, A., Altman, A. 1996.Agrobacterium rhizogenes-mediated DNA transfer inPinus halepensis Mill [J]. Plant Cell Rep., 16: 26–31.Google Scholar
  26. Walter, C., Grace, L.J., Wagner, A., White, D.W.R., Walden, A.R., Donaldson, S.S., Hinton, H., Gardner, R.C., Smith, D.R. 1998. Stable transformation and regeneration of transgenic plants ofPinus radiata D. Don [J]. Plant Cell Rep., 17: 460–469.CrossRefGoogle Scholar
  27. Walter, C., Grace, L., Donaldson, S.S., Moody, J., Gemmell, J.E., van der Maos, S., Kvaalen, H., Lonneborg, A. 1999. An efficient Biolistic transformation protocol forPicea abies embryogenic tissue and regeneration of transgenic plants [J]. Can. J. For. Res., 29: 1539–1546.CrossRefGoogle Scholar
  28. Xiang, Y., Wong, W.K.R., Ma, M.C., Wong, R.S.C. 2000.Agrobacterium-mediated transformation ofBrassica campestris spp. Parachinensis with syntheticBacillus thuringiensis cry1Ab and cry1Ac genes [J]. Plant Cell Rep., 19: 251–256.CrossRefGoogle Scholar

Copyright information

© Northeast Forestry University 2002

Authors and Affiliations

  1. 1.Forest Biotechnology GroupNorth Carolina State UniversityRaleighUSA

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