Testing For Genetically Modified Foods

Polymerase Chain Reaction (PCR) identifies sequences of DNA that have been inserted into the GM plant. In contrast to proteins, DNA is a relatively stable molecule, thus DNA fragments can be isolated from highly processed goods and are sufficiently intact to be amplified by PCR. Genetic engineers use only a small number of regulatory sequences (promoter and terminator sequences) to control the expression of the inserted genes, and so these sequences are common to the majority of GM crops. The two sequences identified in this procedure are two of the most common regulatory sequences, 35S promoter gene from cauliflower mosaic virus (CaMV) and the nopaline synthase (NOS) terminator gene from Agrobacterium tumefaciens.

PCR involves repetitive cycles, each consisting of template denaturation, primer annealing, and extension of the annealed primer by Taq DNA polymerase. Once DNA is extracted from food, a thermal cycler is used to rapidly manipulate temperature, causing the stages of the PCR cycles.

The denaturing stage occurs when samples are rapidly heated to 94 °C, causing the DNA strands to separate. Rapid cooling to 59 °C allows primers to anneal to the separated DNA strands, then reheat to 72 °C for Taq DNA polymerase to extend the primers, making complete copies of each DNA strand and completing one thermal cycle.

The amplified DNA can then be run through an agarose gel with electrophoresis to separate the DNA into visible bands for identification of the 35S promoter gene and the NOS terminator. Amplified DNA is loaded into wells at one end of the gel, using a purchased loading dye that helps weigh down the sample to prevent dissolution into the surrounding buffer. The loading dye also provides a visual, so the movement of DNA can be seen during electrophoresis in the dye “front.” The electrophoresis process works by using an electric current separated into cathode and anode ends. DNA is loaded into the gel on the end closest to the cathode side of the chamber, and the negative charge of DNA is attracted to the anode end of the chamber and is pulled through the agarose. Larger sequences of DNA (increased number of base pairs) cannot move as easily through agarose and will separate out early, while the smaller sequences are able to travel farther down the gel toward the anode end.

A staining process helps by binding to the DNA in order to add contrast between the background gel and the banks of DNA for better visualization of results. Using provided known locations for the different sizes of DNA sequences each test gel can be analyzed for the presence or absence of the 35S promoter and NOS terminator genes.

Controls are used to ensure DNA is extracted properly and to provide a comparison to the test food samples. Purchased plant primer is added to each sample to provide nucleic acids common to all plants. This allows for a quality control check on the DNA extraction process, because any plant DNA extracted should be extended with this primer during PCR and should also be seen on the gel after electrophoresis is complete. Purchased primer for the 35S and NOS genes are used as a positive control to provide DNA bands on the gel for genetic modification. If the GMO-positive template control does not amplify, there is a problem with the PCR reaction and a GMO-negative result from the test food cannot be trusted. A certified non-GMO food product is also purchased and used as a negative control to show what DNA separation looks like when no genetically modified material is present.

After destaining, gels can be analyzed by looking at test food lanes (Table 3) to determine if the DNA bands for the 35S promoter and NOS terminator genes are present in the known locations on the gel. Placing the gel on a UV light box can help provide increased contrast (Figure 1). Alternatively, gels can be placed on white or yellow paper to provide a contrasting background to highlight DNA bands (Figure 2).

Figure 1. A destained gel showing separated bands of DNA. Agarose gel following agarose gel electrophoresis on UV light box.

Figure 2. Diagram of known locations for the 35S promoter and NOS terminator DNA. The presence or absence of a 200 bp band in lane 5 indicates whether or not the test food contains GMOs.

Table 3. PCR Sample Band Sizes (base pairs (bp)).

Polymerase Chain Reaction (PCR) is used to amplify DNA, allowing for a wide range of DNA lab testing. One area of testing now possible with PCR is to identify GMOs by testing for presence or absence of the DNA sequences used in the genetic modification of food crops. Typically, a crop is genetically modified to confer an advantage against natural deterrents to ideal yields, e.g. pests (Figure 3), diseases, drought conditions (Figure 4), etc. Because the advantage is gained by inserting genetic material from a different species into the crop plant’s own DNA, potential human health and environmental risks have been identified with the use of GMOs. One environmental concern is the ability of the genetically modified DNA to be exchanged unintentionally through pollination processes, which could lead to genetically modified DNA entering the genomes of crops intended to be sold as non-GMOs.

Figure 3. Larvae of Colorado beetle, devouring leaves of a potato.

Figure 4. Corn destroyed by drought.