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. 1988 Nov;85(22):8502–8505. doi: 10.1073/pnas.85.22.8502

Stable genetic transformation of intact Nicotiana cells by the particle bombardment process

Theodore M Klein *, Elisabeth C Harper †, Zora Svab †,‡, John C Sanford §, Michael E Fromm *, Pal Maliga †,‡
PMCID: PMC282486  PMID: 16593993

Abstract

We show that the genetic transformation of Nicotiana tabacum can be achieved by bombarding intact cells and tissues with DNA-coated particles. Leaves or suspension culture cells were treated with tungsten microprojectiles carrying plasmid DNA containing a neomycin phosphotransferase gene. Callus harboring the foreign gene was recovered from the bombarded tissue by selection on medium containing kanamycin. Kanamycin-resistant plants have subsequently been regenerated from the callus derived from leaves. Transient expression of an introduced β-glucuronidase gene was used to assess the efficiency of DNA delivery by microprojectiles. The frequency of cells that were stably transformed with the neomycin phosphotransferase gene was a few percent of the cells that transiently expressed the β-glucuronidase gene. These results show that gene transfer by high-velocity microprojectiles is a rapid and direct means for transforming intact plant cells and tissues that eliminates the need for production of protoplasts or infection by Agrobacterium.

Keywords: gene transfer, transgenic plants, microprojectiles, particle gun, Nicotiana tabacum

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Cocking E. C., Davey M. R. Gene transfer in cereals. Science. 1987 Jun 5;236(4806):1259–1262. doi: 10.1126/science.236.4806.1259. [DOI] [PubMed] [Google Scholar]
  2. Czernilofsky A. P., Hain R., Herrera-Estrella L., Lörz H., Goyvaerts E., Baker B. J., Schell J. Fate of selectable marker DNA integrated into the genome of Nicotiana tabacum. DNA. 1986 Apr;5(2):101–113. doi: 10.1089/dna.1986.5.101. [DOI] [PubMed] [Google Scholar]
  3. De Block M., Herrera-Estrella L., Van Montagu M., Schell J., Zambryski P. Expression of foreign genes in regenerated plants and in their progeny. EMBO J. 1984 Aug;3(8):1681–1689. doi: 10.1002/j.1460-2075.1984.tb02032.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. De Greve H., Dhaese P., Seurinck J., Lemmers M., Van Montagu M., Schell J. Nucleotide sequence and transcript map of the Agrobacterium tumefaciens Ti plasmid-encoded octopine synthase gene. J Mol Appl Genet. 1982;1(6):499–511. [PubMed] [Google Scholar]
  5. Fromm M. E., Taylor L. P., Walbot V. Stable transformation of maize after gene transfer by electroporation. 1986 Feb 27-Mar 5Nature. 319(6056):791–793. doi: 10.1038/319791a0. [DOI] [PubMed] [Google Scholar]
  6. Gamborg O. L., Miller R. A., Ojima K. Nutrient requirements of suspension cultures of soybean root cells. Exp Cell Res. 1968 Apr;50(1):151–158. doi: 10.1016/0014-4827(68)90403-5. [DOI] [PubMed] [Google Scholar]
  7. Goodman R. M., Hauptli H., Crossway A., Knauf V. C. Gene transfer in crop improvement. Science. 1987 Apr 3;236(4797):48–54. doi: 10.1126/science.236.4797.48. [DOI] [PubMed] [Google Scholar]
  8. Harpster M. H., Townsend J. A., Jones J. D., Bedbrook J., Dunsmuir P. Relative strengths of the 35S cauliflower mosaic virus, 1', 2', and nopaline synthase promoters in transformed tobacco sugarbeet and oilseed rape callus tissue. Mol Gen Genet. 1988 Apr;212(1):182–190. doi: 10.1007/BF00322463. [DOI] [PubMed] [Google Scholar]
  9. Jefferson R. A., Kavanagh T. A., Bevan M. W. GUS fusions: beta-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J. 1987 Dec 20;6(13):3901–3907. doi: 10.1002/j.1460-2075.1987.tb02730.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Klein T. M., Fromm M., Weissinger A., Tomes D., Schaaf S., Sletten M., Sanford J. C. Transfer of foreign genes into intact maize cells with high-velocity microprojectiles. Proc Natl Acad Sci U S A. 1988 Jun;85(12):4305–4309. doi: 10.1073/pnas.85.12.4305. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Koren G., Lau A., Klein J., Golas C., Bologa-Campeanu M., Soldin S., MacLeod S. M., Prober C. Pharmacokinetics and adverse effects of amphotericin B in infants and children. J Pediatr. 1988 Sep;113(3):559–563. doi: 10.1016/s0022-3476(88)80653-x. [DOI] [PubMed] [Google Scholar]
  12. Paszkowski J., Shillito R. D., Saul M., Mandák V., Hohn T., Hohn B., Potrykus I. Direct gene transfer to plants. EMBO J. 1984 Dec 1;3(12):2717–2722. doi: 10.1002/j.1460-2075.1984.tb02201.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Rhodes C. A., Pierce D. A., Mettler I. J., Mascarenhas D., Detmer J. J. Genetically transformed maize plants from protoplasts. Science. 1988 Apr 8;240(4849):204–207. doi: 10.1126/science.2832947. [DOI] [PubMed] [Google Scholar]
  14. Riggs C. D., Bates G. W. Stable transformation of tobacco by electroporation: evidence for plasmid concatenation. Proc Natl Acad Sci U S A. 1986 Aug;83(15):5602–5606. doi: 10.1073/pnas.83.15.5602. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. St Schell J. Transgenic plants as tools to study the molecular organization of plant genes. Science. 1987 Sep 4;237(4819):1176–1183. doi: 10.1126/science.237.4819.1176. [DOI] [PubMed] [Google Scholar]
  16. Thomashow M. F., Nutter R., Montoya A. L., Gordon M. P., Nester E. W. Integration and organization of Ti plasmid sequences in crown gall tumors. Cell. 1980 Mar;19(3):729–739. doi: 10.1016/s0092-8674(80)80049-3. [DOI] [PubMed] [Google Scholar]
  17. Vieira J., Messing J. Production of single-stranded plasmid DNA. Methods Enzymol. 1987;153:3–11. doi: 10.1016/0076-6879(87)53044-0. [DOI] [PubMed] [Google Scholar]

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