Toward metrological traceability for DNA fragment ratios in GM quantification. 3. Suitability of DNA calibrants studied with a MON 810 corn model.
The quantification of GMOs by real-time PCR relies on an external calibrant. In this paper the suitability of two DNA calibrants, genomic DNA from plant leaves and plasmidic DNA, was investigated. The PCR efficiencies, the correlation coefficients of the calibration curves, and the ratios between PCR efficiencies of transgenic and endogenous sequences were compared for both calibrants using 59 data sets produced by 43 laboratories. There were no significant differences between plasmidic and genomic DNA except for the PCR efficiencies of the calibration curves for the transgene of the construct-specific real-time PCR method. In the GM system investigated, PCR efficiencies of plasmidic calibrants were slightly closer to the PCR efficiencies observed for the unknowns than those of the genomic DNA calibrant. Therefore, plasmidic DNA was the more suitable calibrant for the PCR measurements on genomic DNA extracted from MON 810 seeds. It is shown that plasmidic DNA is an appropriate choice for the calibration of measurements of MON 810 corn with respect to the DNA copy number ratio.
Charels
Diana
D
European Commission, Joint Research Centre, Institute for Reference Materials and Measurements, IRMM, Retieseweg 111, 2440 Geel, Belgium. diana.charels@ec.europa.eu
Broeders
Sylvia
S
Corbisier
Philippe
P
Trapmann
Stefanie
S
Schimmel
Heinz
H
Emons
Hendrik
H
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000235:genetics; D001427:Bacterial Toxins / Q000235:genetics; D002138:Calibration; D018744:DNA, Plant / Q000032:analysis*; D004731:Endotoxins / Q000235:genetics; D030841:Food, Genetically Modified; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics; D006460:Hemolysin Proteins / Q000235:genetics; D018515:Plant Leaves / Q000235:genetics; D030821:Plants, Genetically Modified / Q000235:genetics*; D016133:Polymerase Chain Reaction; D012639:Seeds / Q000235:genetics; D003313:Zea mays / Q000235:genetics*
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D018744:DNA, Plant; D004731:Endotoxins; D018840:HSP70 Heat-Shock Proteins; D006460:Hemolysin Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis