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Evaluation of Plasmid and Genomic DNA Calibrants Used for the Quantification of Genetically Modified Organisms

Evaluation of Plasmid and Genomic DNA Calibrants Used for the Quantification of Genetically Modified Organisms

M Caprioara-Buda, W Meyer, B Jeynov, P Corbisier, S Trapmann, H Emons

Anal Bioanal Chem. 2012 Jul;404(1):29-42.

PMID: 22638881

Abstract:

The reliable quantification of genetically modified organisms (GMOs) by real-time PCR requires, besides thoroughly validated quantitative detection methods, sustainable calibration systems. The latter establishes the anchor points for the measured value and the measurement unit, respectively. In this paper, the suitability of two types of DNA calibrants, i.e. plasmid DNA and genomic DNA extracted from plant leaves, for the certification of the GMO content in reference materials as copy number ratio between two targeted DNA sequences was investigated. The PCR efficiencies and coefficients of determination of the calibration curves as well as the measured copy number ratios for three powder certified reference materials (CRMs), namely ERM-BF415e (NK603 maize), ERM-BF425c (356043 soya), and ERM-BF427c (98140 maize), originally certified for their mass fraction of GMO, were compared for both types of calibrants. In all three systems investigated, the PCR efficiencies of plasmid DNA were slightly closer to the PCR efficiencies observed for the genomic DNA extracted from seed powders rather than those of the genomic DNA extracted from leaves. Although the mean DNA copy number ratios for each CRM overlapped within their uncertainties, the DNA copy number ratios were significantly different using the two types of calibrants. Based on these observations, both plasmid and leaf genomic DNA calibrants would be technically suitable as anchor points for the calibration of the real-time PCR methods applied in this study. However, the most suitable approach to establish a sustainable traceability chain is to fix a reference system based on plasmid DNA.

Chemicals Related in the Paper:

Catalog Number	Product Name	Price
AS2121441	NK603 pDNA calibrant	Price
AS2121442	356043 pDNA calibrant	Price
AS2121443	98140 pDNA calibrant	Price
AS24559	Soya 356043 GMO standard ERM-BF425	Price
AS24567	Maize GMO Standard 98140	Price

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