( content of an unknown sample was then ...)Detection of Genetically Modified Soybean in Processed Foods Using Real-Time Quantitative PCR with SYBR Green I Dye on the DNA Engine Opticon 2 System

content of an unknown sample was then calculated from the Trend line of the ΔC(t) values for the known GMO concentration standards. As can be seen in Table 4, the soy dessert was found to contain 4.4% GMO soy. Since the ΔC(t) value for the soy burger was outside the range of values for the standards (more than that of the 5% standard), the GMO content was calculated from the trend as ~10.32%. Note that for both food samples the amount of GMO soy being calculated using the ΔC(t) value represents the amount of GMO soy relative to the total amount of soy present in the sample.

The results of the melting curve analysis are presented in Figures 2 and 3. Plotting the negative first derivative shows the melting point (T_m) as a single peak at approximately 88C for reactions with the lectin primers and 92C for reactions with EPSPS primers (Figure 2). For all DNA samples, a single peak at 88C confirmed the presence of soy (Table 5). Among the food samples tested, only the DNA from the burger and soy dessert samples generated a distinct peak at 92C, consistent with the presence of GMO soy (Figure 2 and Table 4).

To eliminate the possibility that the soy flour samples failed to generate the epsps amplicon due to an inhibition of the PCR by some component of the isolated DNA, control reactions with internal standards were run in parallel with the samples being analyzed. Each food sample was spiked with 20mg of the 5% GMO reference standard. DNA was then isolated from the spiked sample and amplified in reactions with the lectin primers and the EPSPS primers. Comparison of the melting curves of the spiked and nonspiked soy flour samples show that the EPSPS-specific pe
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