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Technical Abstract Details

Title 037-02 Evaluation of DNA Recovery Methods Using Real-Time PCR for the Detection of Soy in Foods
Presenter Chern Lin Koh, Christina A Mireles DeWitt, Oregon State Univ., Corvallis, OR
Abstract Under the Food Allergen Labeling and Consumer Protection Act, foods that contain or derive from the Big 8 allergens must be declared and printed on the label of the food product. Since soybean products are highly processed, ELISA-based detection method for soy allergen can give incorrect results. Thus, the objective of this study was to investigate the application of Wizard Magnetic DNA Purification system kit (Promega, Madison, Wis.) and DNeasy mericon Food Kit (Qiagen, Valencia, Calif.) for soybean detection using Real-Time PCR. Both protocols were conducted using 150-200 mg of soy isolate protein and powdered soybean. The extracted DNA was diluted with deionized water (ranging from 2,000 ppm to 0.1 ppb) prior to purification to determine the detection limits. Extractions were conducted in triplicate and DNA was collected. DNA recovered from extraction was stored at -20 ºC. Real-Time PCR was performed with primers designed for the lectin gene (Le1) in soybean and standard curves were constructed. Pisum sativum (Green pea) was extracted with both kits, diluted with deionized water (ranged from 100,000 to 10 ppm), and used as negative control to verify the specificity of the primer. In terms of primer verification, no response was detected for the negative control. For samples extracted with DNeasy mericon Food Kit, the response showed an average R2 value of 0.99 with detection ranging from 2,000 to 2 ppm. However, the Wizard Magnetic DNA Purification system showed a nonlinear and random response ranging from 1,000 ppm to 1 ppb for isolated protein and 1,000 to 1 ppm for powdered soybean. Such observation could be linked to the binding affinity, surface interaction, and limitation of the magnetic beads. Subsequent research will evaluate both protocols against ELISA-based method in food products.
Year/Location2012 IFT Annual Meeting, June 25 - 28, Las Vegas NV
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