Validated Method for Quantification of Gentically Modified Organisms in Samples of Maize Flour

Institute of Applied Microbiology, University of Natural Resources and Applied Life Sciences, Muthgasse 18, A-1190 Vienna, Austria.
Journal of Agricultural and Food Chemistry (Impact Factor: 2.91). 03/2006; 54(3):678-81. DOI: 10.1021/jf052257s
Source: PubMed


Sensitive and accurate testing for trace amounts of biotechnology-derived DNA from plant material is the prerequisite for detection of 1% or 0.5% genetically modified ingredients in food products or raw materials thereof. Compared to ELISA detection of expressed proteins, real-time PCR (RT-PCR) amplification has easier sample preparation and detection limits are lower. Of the different methods of DNA preparation CTAB method with high flexibility in starting material and generation of sufficient DNA with relevant quality was chosen. Previous RT-PCR data generated with the SYBR green detection method showed that the method is highly sensitive to sample matrices and genomic DNA content influencing the interpretation of results. Therefore, this paper describes a real-time DNA quantification based on the TaqMan probe method, indicating high accuracy and sensitivity with detection limits of lower than 18 copies per sample applicable and comparable to highly purified plasmid standards as well as complex matrices of genomic DNA samples. The results were evaluated with ValiData for homology of variance, linearity, accuracy of the standard curve, and standard deviation.

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    • "Hellemans et al. [41] advise to make the dilution series with a sample that mimics as much as possible the samples to be analysed in qPCR [41], most often this is a mixture of representative cDNA samples [57,75]. Plasmid DNA consists of a different sample matrix, what can result in altered efficiencies due to the presence of different kinds of inhibitory components [76]. However, the absence of PCR inhibitors was controlled for by means of the SPUD assay. "
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