Abstract
Stable genetic transformation ofPicea mariana (black spruce) was obtained via particle bombardment into two target tissues, mature cotyledonary somatic embryos and suspensions from embryonal masses, with the Biolistic PDS-1000/He device. Seven transgenic embryogenic cell line were obtained from the mature cotyledonary somatic embryos after secondary somatic embryogenesis from two different cell lines (R4F14 and 119794-014). The suspension culture from embryonal masses produced five transgenic cell lines from one cell line (R4F14). Expression of the introduced β-glucuronidase (GUS) and neomycin phosphotransferase II (NPT II) genes was detected by histochemistry and fluorometry, and by ELISA in 10 of the lines. Two lines showed only NPT II gene expression. Four of the five lines obtained after bombardment of suspensions of embryonal masses showed lower levels of expression of GUS and NPT II. The integration of the foreign genes was confirmed by polymerase chain reaction analyses and Southern hybridization for GUS and NPT II, and complex hybridization patterns were observed. The 12 transgenic lines obtained had a typical embryogenic morphology and were capable of maturation and germination. Over 40 transgenic trees were regenerated from one of the transgenic lines, and they have a normal phenotype.
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Barton, K. A.; Binns, A. N.; Matzke, A. J. M., et al. Regeneration of intact tobacco plants containing full length copies of genetically engineered T-DNA, and transmission of T-DNA to R1 progeny. Cell 32:1033–1043; 1983.
Bommineni, V. R.; Chibbar, R. N.; Datla, R. S. S., et al. Transformation of white spruce (Picea glauca) somatic embryos by microprojectile bombardment. Plant Cell Rep. 13:17–23; 1993.
Braun, C. F.; Jilka, J. M.; Hemenway, C. L., et al. Interactions between plants, pathogens and insects: possibilities for engineering resistance. Curr. Opin. Biotech. 2:193–198; 1991.
Carlson, J. E.; Tulsieram, L. K.; Glaubitz, J. C., et al. Segregation of random amplified DNA markers in F1 progeny of conifers. Theor. Appl. Genet. 83:194–200; 1991.
Castillo, A. M.; Vasil, V.; Vasil, I. K. Rapid production of fertile transgenic plants of rye (Secale cereale L.). Bio Technology 12:1366–1371; 1994.
Charest, P. J.; Calero, N.; Lachance, D., et al. The use of microprojectile DNA delivery to bypass the long life cycle of tree species in gene expression studies. Curr. Top. Bot. Res. 1:151–163; 1993a.
Charest, P. J.; Calero, N.; Lachance, D., et al. Microprojectile DNA delivery in conifer species: factors affecting assessment of transient gene expression using the β-glucuronidase reporter gene. Plant Cell Rep. 12:189–193; 1993b.
Charest, P. J.; Michel, M. F. Basics of plant genetic engineering and potential applications to tree species. Petawawa National Forestry Institute, Information Report PI-X-104. 1991:48p.
Cheliak, W. M.; Klimaszewska, K. K. Genetic variation in somatic embryogenic response in open-pollinated families of black spruce. Theor. Appl. Genet. 82:185–190; 1991.
Christou, P.; Ford, T. L.; 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. BioTechnology 9:957–962; 1991.
Datla, R. S. S.; Bekkaoui, F.; Hammerlindl, J. K., et al. Improved high-level constitute foreign gene expression in plants using an AMV RNA4 untranslated leader sequence. Plant Sci. 94:139–149; 1993.
Datla, R. S. S.; Hammerlindl, J. K.; Pelcher, L. E., et al. A bifunctional fusion between β-glucuronidase and neomycin phosphotransferase: a broad-spectrum marker enzyme for plants. Gene 101:239–246; 1991.
Ellis, D. D.; McCabe, D. E.; McInnis, S., et al. Stable transformation ofPicea glauca by particle acceleration. Bio Technology 11:84–89; 1993.
Gordon-Kamm, W. J.; Spencer, M.; Mangano, M. L., et al. Transformation of maize cells and regeneration of fertile transgenic plants. Bio-Technology 2:603–618; 1990.
Herrera-Estrella, L.; Depicker, A.; van Montagu, M., et al. Expression of chimeric genes transferred into plant cells using a Ti-plasmid-derived vector. Nature 303:209–213; 1983.
Horsch, R. B.; Fry, J. E.; Hoffman, N., et al. A simple and general method for transferring genes into plants. Science 227:1129–1132; 1985.
Huang, Y.; Diner, A. M.; Karnosky, D. F..Agrobacterium rhizogenes-mediated genetic transformation and regeneration of a conifer:Larix decidua. In Vitro Cell Dev. Biol. 27:201–207; 1991.
Jefferson, R. A.; Kavanagh, T. A.; Bevan, M. W.. GUS fusion: β-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J. 6:3901–3907; 1987.
Kay, R.; Chan, A.; Daly, M., et al. Duplication of CaMV 35S promoter sequences creates a strong enhancer for plant genes. Science 236: 1299–1302; 1987.
Klein, T. M.; Wolf, E. D.; Wu, R., et al. High-velocity microprojectile for delivering nucleic acids into living cells. Nature 327:70–73; 1987.
Lelu, M. A.; Klimaszewska, K.; Jones, C. et al. A laboratory guide to somatic embryogenesis in spruce and larch. Petawawa National Forestry Institute, Information Report PI-X-111, 1993;57p.
Maniatis, T.; Fritsch, E. F.; Sambrook, J. Molecular cloning—a laboratory manual. Cold Spring Harbor, New York: Cold Springer Harbor Laboratory Press; 1982:545p.
Raman, K. V.; Altman, D. W. Biotechnology initiative to achieve plant pest and disease resistance. Crop Protetion 13:591–596; 1994.
Register, J. C., III; Peterson, D. J.; Bell, P. J., et al. Structure and function of selectable and non-selectable transgenes in maize after introduction by particle bombardment. Plant Mol. Biol. 25:951–961; 1994.
Robertson, D.; Weissinger, A. K.; Ackley, R., et al. Genetic transformation of Norway spruce (Picea abies L.) Karst using somatic embryo explants by microprojectile bombardment. Plant Mol. Biol. 19:925–935; 1992.
Seguin, A.; Lachance, D.; Charest, P. J. Transien gene expression and stable genetic transformation into conifer tissues by microprojectile bombardment. Plant Tissue Cult. Man. (In press).
Somers, D. A.; Rines, H. W.; Gu, W., et al. Fertile, transgenic oat plants. BioTechnology 10:1589–1594; 1992.
Tomes, D. T.; Weissinger, A. K.; Ross, M., et al. Transgenic tobacco plants and their progeny derived by microprojectile bombardment of tobacco leaves. Plant Mol. Biol. 14:261–268; 1990.
Topfer, R.; Schell, J.; Steinbiss, H.-H. Versatile cloning vectors for transient gene expression and direct gene transfer in plant cells. Nucleic Acid Res. 16:8725; 1988.
Vasil, V.; Castillo, A. M.; Fromm, M. E., et al. Herbicide resistant fertile transgenic wheat plants obtained by microprojectile bombardment of regenerable embryogenic callus. BioTechnology 10:667–674; 1992.
von Aderkas, P.; Bonga, J.; Klimazewska, K., et al. Comparison of larch embryogeny in vivo and in vitro. In: Ahuja, M. R., ed. Woody plant biotechnology. New York: Plenum Press; 1991:139–155.
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Charest, P.J., Devantier, Y. & Lachance, D. Stable genetic transformation ofPicea mariana (black spruce) via particle bombardment. In Vitro Cell.Dev.Biol.-Plant 32, 91–99 (1996). https://doi.org/10.1007/BF02823137
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DOI: https://doi.org/10.1007/BF02823137