Abstract
Legislation in the EU requires that foods containing more than 0.9% of genetically modified organisms (GMOs) should be labelled. To this end, we have developed a simple and accurate capillary electrophoresis multiplex quantitative competitive PCR (ce-mqcPCR) method for event-specific quantification of the five novel GM maize events DAS59122, LY038, MON88017, MIR604 and Event 3272. The method combines the simplicity of constructing multiple competitors in silico with the high resolution and sensitivity of fluorescence capillary electrophoresis and the use of an internal template reference amplicon. The competitors are synthesised commercially and added in equal amounts as a restriction enzyme-digested plasmid insert to the multiplex PCR. Quantification is performed by analysing the relative amounts of GMO and GMO competitor fragment pairs after capillary electrophoresis and correcting for differences in maize DNA by comparing with the internal reference gene amplicon. Since the competitors employ the same primers as their corresponding targets, all existing qualitative multiplex PCRs can in principle easily be converted to quantitative assays without changing primer sets or amplification conditions. The ce-mqcPCR method correctly determined 120 GMO templates in known mixed samples. No false-positive or false-negative signals were obtained.
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Acknowledgments
We thank Dr. P. Lea, Nofima AS, Norway, for statistical analyses. The authors declare that they have no conflict of interest. This work was supported by the Norwegian Research Council project 154254/130 and the European Commission through the Integrated Project Co-Extra, Contract No. 007158, under the 6th Framework Programme, priority 5, food quality and safety.
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Holck, A.L., Pedersen, B.O. Simple, sensitive, accurate multiplex quantitative competitive PCR with capillary electrophoresis detection for the determination of genetically modified maize. Eur Food Res Technol 233, 951–961 (2011). https://doi.org/10.1007/s00217-011-1594-6
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DOI: https://doi.org/10.1007/s00217-011-1594-6