Advertisement

European Food Research and Technology

, Volume 216, Issue 4, pp 347–354 | Cite as

Event specific real-time quantitative PCR for genetically modified Bt11 maize (Zea mays)

  • Sissel B. Rønning
  • Marc Vaïtilingom
  • Knut G. Berdal
  • Arne Holst-JensenEmail author
Original Paper

Abstract.

We report the characterisation of the 3'-integration junction between host plant DNA and integrated gene-construct of the genetically modified Bt11 maize, and the successive development and first validation of a transformation event-specific quantitative TaqMan 5'-nuclease PCR assay for detection and quantitation of Bt11 maize on the LightCycler. The transgenic DNA in Bt11 is inserted in a tandem repeated DNA sequence motif of approximately 180 bp, and we discuss some of the problems this creates for design, development and validation of event-specific PCR assays. The limit of detection of the PCR assay was estimated to 10 initial template copies. The limit of quantitation in pure maize materials was estimated to approximately 40 template copies. In food samples containing high levels of exogenous DNA, the impact of this DNA was estimated to increase the limit of quantitation to approximately 100 initial template copies. Because of the low number of template copies required for detection and quantification, in combination with small size of the amplicon (~100 bp), the assay may be applied to analyses of raw materials as well as processed food and feed.

Keywords

Event-specific PCR Real-time PCR Genetically modified organism GMO Bt11 maize Detection and quantitation limit Integration site 

Notes

Acknowledgements

We wish to thank Martha Hernandez Perez, CSIC, Barcelona, Spain for providing 100% Bt11 material. This study was financially supported by the EC FP5 "Quality of Life and Management of Living Resources" program within the project "Reliable, standardised, specific, quantitative detection of genetically modified food" (contract no. QLK1–1999–01301, acronym Qpcrgmofood), and by a grant from the Research Council of Norway (136430/140). This is gratefully acknowledged.

Reference List

  1. 1.
    James C (2001) ISAAA Briefs No. 24Google Scholar
  2. 2.
    Regulation EC (1997) Official Journal of the European Communities No. L 43:1–5Google Scholar
  3. 3.
    Regulation EC (1998) Official Journal of the European Communities No. L 159/4:4–7Google Scholar
  4. 4.
    Commission Regulation EC (2000) Official Journal of the European Communities No. L6:13–14Google Scholar
  5. 5.
    Commission Regulation EC (2000) Official Journal of the European Communities No. L 6:15–17Google Scholar
  6. 6.
    Anklam E, Gadani F, Heinze P, Pijnenburg H, Van den Eede G (2002) Eur Food Res Technol 214:3–26Google Scholar
  7. 7.
    Wolf C, Scherzinger M, Wurz A, Pauli U, Hubner P, Luthy J (2000) Eur Food Res Technol 210:367–372Google Scholar
  8. 8.
    Agbios database (2002) http://www.agbios.com/default.aspGoogle Scholar
  9. 9.
    Berdal KG, Holst-Jensen A (2001) Eur Food Res Technol 213:432–438Google Scholar
  10. 10.
    Terry CF, Harris N (2001) Eur Food Res Technol 213:425–431Google Scholar
  11. 11.
    Taverniers I, Windels P, Van Bockstaele E, De Loose M (2001) Eur Food Res Technol 213:417–424Google Scholar
  12. 12.
    Holck A, Vaïtilingom M, Didierjean L, Rudi K (2002) Eur Food Res Technol 214:449–454Google Scholar
  13. 13.
    Zimmermann A, Luthy J, Pauli U (2000) Lebensm Wiss Technol 33:210–216Google Scholar
  14. 14.
    Vaïtilingom M, Pijnenburg H, Gendre F, Brignon P (1999) J Agric Food Chem 47:5261–5266PubMedGoogle Scholar
  15. 15.
    Wurz A, Bluth A, Zeltz P, Pfeifer C, Willmund R (1999) Food Contr 10:385–389CrossRefGoogle Scholar
  16. 16.
    Heid CA, Stevens J, Livak KJ, Williams PM (1996) Genome Res 6:986–994PubMedGoogle Scholar
  17. 17.
    Hartl D, Ochman H (1996) Meth Mol Biol 58:293–301Google Scholar
  18. 18.
    Van den Eede G, Lipp M, Eyquem F, Anklam E (2000) Report from the EC-JRC, Ispra, Italy, EUR 19677Google Scholar
  19. 19.
    Landt O, Peters M (1998) http://www.lynet.de/TIB-MOLBIOL/oligo_ag.htmlGoogle Scholar
  20. 20.
    Xu J, Pemberton GH, Almira EC, McCarty DR, Koch KE (1995) Plant Physiol 108:1293–1294CrossRefPubMedGoogle Scholar
  21. 21.
    Arumuganathan K, Earle ED (1991) Plant Mol Biol Rep 9:211–215Google Scholar
  22. 22.
    Dennis ES, Peacock WJ (1984) J Mol Evol 20:341–350PubMedGoogle Scholar

Copyright information

© Springer-Verlag 2003

Authors and Affiliations

  • Sissel B. Rønning
    • 1
  • Marc Vaïtilingom
    • 2
  • Knut G. Berdal
    • 1
  • Arne Holst-Jensen
    • 1
    Email author
  1. 1.Section of Food and Feed MicrobiologyNational Veterinary InstituteOsloNorway
  2. 2.TEPRALStrasbourgFrance

Personalised recommendations