Skip to main content
Log in

Transgene copy number can be positively or negatively associated with transgene expression

  • Research Article
  • Published:
Plant Molecular Biology Aims and scope Submit manuscript

Abstract

Two different types of T-DNA insert were found in tobacco plants transformed with Agrobacterium tumefaciens. High-expressing (H) types had one copy of the T-DNA at a locus and produced high expression of the transgene uidA, as measured by uidA RNA levels and β-glucuronidase activity; low-expressing (L) types had inverted repeats of the T-DNA at a locus and produced low uidA expression. H-types from different transformants acted additively, and cross-fertilization between two different homozygous transformants with H-type inserts produced F1 plants with GUS activity that equalled the parents and individual F2 plants with 50%, 100%, 150% and 200% of parental values. However, the L-type inserts worked in trans to suppress uidA expression from H-type inserts when both were present in the same genome. Hence when a transformant homozygous for the L-type insert was crossed to one homozygous for the H-type, all plants in the F1 and F2 generations with both types of insert had low GUS activity while F2 segregants that only had the H-type inserts had high GUS activity again. Suppression of the H-type gene was associated with increased methylation of the insert. Particle acceleration was used to introduce further copies of uidA into tissues of the transformants. Regardless of the promoter used, those plants with endogenous L-type inserts showed none of the distinct loci of GUS activity readily visible in material with no inserts, showing that L-type inserts could suppress not only the uidA expression of genomic homologues, but also of copies added in vitro.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. Bollman J, Carpenter R, Coen ES: Allelic interactions at the nivea locus of Antirrhinum. Plant Cell 3: 1327–1336 (1991).

    Article  PubMed  Google Scholar 

  2. Dean C, Jones J, Favreau M, Dunsmuir P, Bedbrook J: Influence of flanking sequences on variability in expression levels of an introduced gene in transgenic tobacco plants. Nucl Acids Res 16: 9267–9283 (1988).

    PubMed  Google Scholar 

  3. Gasser CS, Fraley RT: Genetically engineering plants for crop improvement. Science 244: 1293–1299 (1989).

    Google Scholar 

  4. Gendloff EH, Bowen B, Buchholz WG: Quantitation of chloramphenicol acetyl transferase in transgenic tobacco plants by EI ISA and correlation with gene copy number. Plant Mol Biol 14: 575–583 (1990).

    Article  PubMed  Google Scholar 

  5. Goring DR, Thomson L, Rothstein SJ: Transformation of a partial nopaline synthase gene into tobacco suppresses the expression of a resident wild-type gene. Proc Natl Acad Sci USA 88: 1770–1774 (1991).

    PubMed  Google Scholar 

  6. Hobbs SLA, Jackson JA, Baliski DS, DeLong CMO, Mahon JD: Genotype-and promoter-induced variability in transient β-glucuronidase expression in pea protoplasts. Plant Cell Rep 9: 17–20 (1990).

    Article  Google Scholar 

  7. Hobbs SLA, Kpodar P, DeLong CMO: The effect of T-DNA copy number, position and methylation on reporter gene expression in tobacco transformants. Plant Mol Biol 15: 851–864 (1990).

    PubMed  Google Scholar 

  8. Jefferson RA: Assaying chimeric genes in plants: the GUS gene fusion system. Plant Mol Biol Rep 5: 387–405 (1987).

    Google Scholar 

  9. Jones JDG, Gilbert DE, Grady KL, Jorgensen RA: T-DNA structure and gene expression in petunia plants transformed by Agrobacterium tumefaciens C58 derivatives. Mol Gen Genet 207: 478–485 (1987).

    Article  Google Scholar 

  10. Jorgensen R: Altered gene expression in plants due to trans interactions between homologous genes. Trends Biotechnol 8: 340–344 (1990).

    Article  PubMed  Google Scholar 

  11. Koes RE, van Blockland R, Quattrocchio F, van Tunen AJ, Mol JMN: Chalcone synthase promoters in petunia are active in pigmented and unpigmented cell types. Plant Cell 2: 379–392 (1990).

    Article  PubMed  Google Scholar 

  12. Matzke MA, Primig M, Trnovsky J, Matzke AJM: Reversible methylation and inactivation of marker genes in sequentially transformed tobacco plants. EMBO J 8: 643–649 (1989).

    Google Scholar 

  13. Murashige T, Skoog F: A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiol Plant 15: 473–497 (1962).

    Google Scholar 

  14. Napoli C, Lemieux C, Jorgensen R: Introduction of a chimeric chalcone synthase gene into petunia results in reversible co-suppression of homologous genes in trans. Plant Cell 2: 279–289 (1990).

    Article  PubMed  Google Scholar 

  15. Rogers SG, O'Connell K, Horsch RB, Fraley RT: Investigations of factors involved in foreign protein expression in transformed plants. In: Zaitlin M, Day P, Hollaender A (eds) Biotechnology in Plant Science, pp. 219–226. Academic Press, London (1985).

    Google Scholar 

  16. Shirsat AH, Wilford N, Croy RRD: Gene copy number and levels of expression in transgenic plants of a seed specific gene. Plant Sci 61: 75–80 (1989).

    Article  Google Scholar 

  17. Smith CJS, Watson CF, Bird CR, Ray J, Schuch W, Grierson D: Expression of a truncated tomato polygalacturonase gene inhibits expression of the endogenous gene in transgenic plants. Mol Gen Genet 224: 477–481 (1990).

    Article  PubMed  Google Scholar 

  18. van der Krol AR, Mur LA, Beld M, Mol JNM, Stuitje AR: Flavonoid genes in petunia: addition of a limited number of gene copies may lead to suppression of gene expression. Plant Cell 2: 291–299 (1990).

    Article  PubMed  Google Scholar 

  19. Weising K, Schell J, Kahl G: Foreign genes in plants: transfer, structure, expression and applications. Annu Rev Genet 22: 421–477 (1988).

    Article  PubMed  Google Scholar 

  20. Wernsman EA, Matzinger DF: Tobacco. In: Fehr WR, Hadley HH (eds) Hybridization of Crop Plants, pp. 657–668. American Society of Agronomy, Madison, WI (1980).

    Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Rights and permissions

Reprints and permissions

About this article

Cite this article

Hobbs, S.L.A., Warkentin, T.D. & DeLong, C.M.O. Transgene copy number can be positively or negatively associated with transgene expression. Plant Mol Biol 21, 17–26 (1993). https://doi.org/10.1007/BF00039614

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00039614

Key words

Navigation