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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

Roundup Ready soybean powders. The following foods were obtained from a supermarket for testing: soy burger, a soy dessert and two brands of soy flour.

The dry samples, the reference standards and the soy flour were used directly in the DNA isolation while wet samples such as the soy burger and the soy dessert were mashed to a fine, homogeneous paste with a mortar and pestle. For each standard and sample, the amount of starting material used for DNA isolation is shown in Table 3.

The DNeasy Plant Mini Kit (Qiagen, 69104) was used for isolation of DNA from reference standards and food samples. DNA was extracted from the different samples as described in the kit protocol. DNA concentration was determined by spectrophotometry and the quality verified by agarose gel electrophoresis on a 1% TBE gel. On average, we were able to obtain between 12g of DNA from each sample material.

Reactions
DyNAzyme II DNA polymerase* was from Finnzymes (F-503L) and SGI was from Molecular Probes (P-7589). Reaction components were assembled in low-profile microplates (MJ Research, MLL-9651) and sealed with ultra-clear strip caps (MJ Research, TCS-0803). Volumes of individual components and final reaction concentrations are listed in Table 1.

SGI was obtained at 10,000X concentration and was diluted to a 10X working concentration with 0.1X TE buffer, pH 8.0. A 10X SGI solution is defined as one giving 0.400.01 OD when measured at 495nm.

Primers for soy lectin and EPSPS were ordered from Sigma Genosys (The Woodlands, Texas). The primers used for amplification of the soy lectin gene generated a 318bp amplicon. Sequences of the lectin primers were: forward, 5-GAC-GCT-ATT-GTG-ACC-TCC-T
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