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Usage Of Pcr To Detect Genetically Modified Food Biology Essay

This paper describes how polymerase chain reaction (PCR) can be used to determine if various food items have been genetically modified by detecting reporter genes. This study involved preparation of 1.25% agarose gel, and the preparation of the various food items for PCR. The food items analyzed in this study were chosen from fresh and processed food items to determine if PCR can detect genetic modification equally. Two different master mixes were used to determine genetic modification. Green master mix coded for the presence of the genetically modified reporter gene GUS, while Red master mix coded for the presence of the genetically modified reporter gene SP 30. PCR was performed to amplify these reporter genes if they were present, and then the samples were analyzed with gel electrophoresis. A digital picture of the gel was taken for analysis. The resulting digital picture of the gel showed that some food items did indeed posses the reporter genes and thus had been genetically modified, while other food items did not posses reporter genes, and thus would appear to have no genetic modification. The results clearly support the importance of PCR in biotechnology applications, in this case the detection of genetic modification of foods.

Introduction:

The term GM foods or GMOs (genetically-modified organisms) refers to crop plants that are created for human or animal consumption using biotechnology techniques. These plants have been modified to enhance desired traits such as increased resistance to herbicides or improved nutritional content. (Campbell, et al. 2005). The traditional method of acquiring the desired traits in plants, ahs bee through breeding, but this can be very time consuming and are often not very accurate. A more accurate method is to genetically engineer the plants, but identifying and changing the specific gene that codes for the desired trait. For example, scientists can isolate a gene that is responsible for producing fruit that is resistant to bug invasion and insert that gene into a different plant that already produces very large, sweet fruit. The new genetically-modified plant will gain the resistance to bug invasion and also retain the production of large, sweet fruit. Organisms that are genetically modified in this way are called transgenic organisms. (Alberts, et al. 2009).

There is a great deal of controversy regarding GM foods, with the proponents of GM foods listing the advantages, while the opponents list the disadvantages of GM foods. Some of the advantages of GM foods are that they can be useful in meeting the world’s food demands due to increased pest resistance, disease resistance, cold tolerance, drought tolerance, and increased nutritional value. Some of the disadvantages of GM foods are that they could cause unintended harm to other organisms based on a gene transfer to another non-target species, which no one knows the results of, but some possibilities are an increase in human health risks due to allergies. (Nordlee et al. 1996).

Along with the advantages and disadvantages are the regulatory issues surrounding GM foods. The world governments all hold different standards for regulating GM foods. Much progress is being made in creating common regulations. (Helmuth. 2000) Regardless of the regulation issues, GM foods have the potential to solve world hunger and other serious issues such as malnutrition. Most people don’t realize that much of the food that we eat in the United States is actually GM foods.

GM foods can be tested for using a variety of methods; however the focus of this paper is on the detection of GM foods by usage of Polymerase Chain Reaction (PCR). When GM foods are created, a reporter gene is encoded into their genome, so that it can easily be determined if the plants DNA has been genetically modified. PCR is simply a process by which you use a primer that specify a particular gene and then amplify it for further study (Bharathan. 2009). In the case of GM foods, PCR can be used to determine if a food is genetically modified or not, by using primers that target the reporter genes that are part of GM foods, and if the reporter gene is present, amplifying them so that they can be detected. The purpose of this paper is to demonstrate how PCR is used in the detection and amplification of reporter genes in GM foods

Materials and Methods:

Two fresh food items, and two processed food items were obtained for testing. The two fresh foods used were Tomato and Green Pepper. The two processed foods used were Tortilla chips and Roasted Peanuts. Approximately 2 grams of each food sample was weighed out and then ground with 10 ml of distilled H2O, using a mortar and pestle, creating a slurry. 50 ul of each sample slurry was placed into a labeled micro centrifuge tube. DNA extraction buffer was provided by Dr. N. Bharathan, and 500 ul of this DNA extraction buffer was added to each of the samples in the micro centrifuge tubes. The contents of each tube was gently mixed, and then placed into hot water incubation @ 95⁰C for 5 minutes. When the incubation period was finished, each of the samples was centrifuged @ 12,000 rpm for 5 minutes.

The samples were then prepared for the PCR reaction. Two micro centrifuge tubes were labeled for each sample. 15 ul of each sample was placed into each of the two corresponding tubes. 15 ul of Green Master Mix, which is a plant primer and codes for the reporter gene GUS, was added to one sample, and 15 ul of Red Master Mix, which is a GMO primer and codes for the reporter gene SP 30, was added to the other sample. The Green and Red Master Mixes were provided by Dr. N. Bharathan. All of the samples were treated in this way, producing two tubes for each sample, one with Red Master Mix added, and one with Green Master Mix added to it. The micro centrifuge tubes were centrifuged momentarily and then placed into a Bio-Rad DNA thermal cycler. The thermal cycler was programmed as follows:

Denature

94C

2 min

X 1 cycle

Denature

94C

1 min

Anneal

59C

1 min

Extend

72C

2 min

X 40 cycles

Extend

72C

10 min

X 1 cycle

Hold

4C

When the thermal cycler reaction was finished, 15 ul of orange loading gel dye was added to each tube to stop the reaction. Gel electrophoresis analysis was conducted using a 1.25% agarose gel.

Results:

The results as seen in Figure 1 show that the samples of Tortilla Chips, Papaya, and the positive control all indicate positive for genetic modification.

GMO group 9.jpg

Figure 1. Digital photograph of Gel after Electrophoresis, showing which food samples tested positive or negative for genetic modification.

Table 1. Data showing positive or negative results for various fresh and processed food items, based on the presence or absence of the reporter genes GUS and SP 30.

Food

Fresh/Processed

SP30

GUS

Plum tomato

Fresh

neg

neg

banana

Fresh

neg

neg

romaine lettuce

Fresh

neg

neg

Strawberries

Fresh

neg

neg

Fresh Corn

Fresh

neg

neg

Potato

Fresh

neg

neg

Peanuts

Processed

neg

neg

cereal

Processed

neg

neg

chip

Processed

neg

neg

green bean

Fresh

neg

neg

Potato Chips

Processed

neg

neg

Grape Tomato

Fresh

neg

neg

Green Peppers

Fresh

neg

neg

Peanuts

Processed

neg

neg

Spinach

Fresh

neg

neg

Corn

Fresh

neg

neg

Strawberry leaves

Fresh

neg

neg

Fruity pebbles

Processed

Pos

neg

Tortilla chips

Processed

Pos

neg

baked lays

Processed

pos

neg

Cin. Toast crunch cereal

Processed

pos

neg

Broccoli

Fresh

Pos

neg

Tostitos Tortilla Chips

Processed

Pos

neg

Salt and Vinegar Potato Chips

Processed

Pos

neg

Peas

Fresh

Pos

neg

Fruity pebbles

Processed

Pos

neg

tomato

Fresh

Pos

neg

peas

Fresh

Pos

neg

carrot

Fresh

Pos

neg

Frosted Flakes

Processed

Pos

neg

Potato Chips

Processed

Pos

neg

papaya

Fresh

pos

pos

Papaya

Fresh

Pos

Pos

Positive Control

 

Pos

Pos

tortilla chips

Processed

pos

pos

Oats

Processed

Pos

Pos

Papaya

Fresh

Pos

Pos

fruity pebbles

Processed

Pos

 

tortilla chips

Processed

Pos

 

Frosted Flakes

Processed

Pos

 

Green Peppers

Fresh

Pos

 

Chinese Yu Choy(Leafy Vege)

Fresh

Pos

 

Tortilla Chips

Processed

Pos

 

Instants Soy Milk Mix

Processed

Pos

 

Discussion:

Genetically modified foods can be detected via the usage of Polymerase Chain Reaction and Gel Electrophoresis. PCR is used to amplify any reporter genes that are present in foods, and that indicate that they have been genetically modified. It is clear from the gel that only Tortilla chips and Papaya contain a reporter gene that indicates that these food items have been genetically modified in some manner. The other food items do not have bands for either the green or the red, and thus are not indicating the presence of any reporter gene in those food items.

Positive and negative controls indicate that both reporter genes should indicate positive to confirm the presence of genetic modification, and this is confirmed by the data from the gel picture. (Figure 1.) However, according to the class data it seems that it is possible that there could be a food item that was positive for only one of the reporter genes. (Table 1). It appears that fewer fresh foods tested positive for either of the reporter genes than did the processed foods (Table 1). This may be attributed to the possibility that food processing companies may choose to use GM foods due to some genetic modification such as having a longer shelf life, or maintaining taste better after being processed.

In conclusion, it is evident that PCR is an efficient and important method that can be used to detect the presence of genetically modification in foods, particularly processed foods. It is also evident that many foods are actually genetically modified. (Table 1).

Literature Cited:

Alberts, B., Bray, D., Hopkin, K., Johnson, A., Lewis, J., Raff, M., Roberts, K., and Walter, P. (2009). Essential Cell Biology (3rd ed. ). New York: Garland Science, Taylor & Francis Group, LLC.

Bharathan, N. 2009. Biology 401: Amplification: The Polymerase Chain Reaction (PCR). Laboratory Techniques in Molecular Biology. Indiana (PA): Indiana University of Pennsylvania. 8 p.

Campbell, N. A., and Reece, J. B. (2005). Biology ( 7th ed. ). New York: Pearson Benjamin Cummings. .

Nordlee, J. A., Taylor, S.L., Townsend, J.A., Thomas, L.A., and Bush, R.K., Identification of a Brazil-nut allergen in transgenic soybeans (New England Journal of Medicine, Vol 334, No 11, pp 688-692, 1996)

Helmuth, L. 2000. Both Sides Claim Victory in Trade Pact. Science. Vol. 287. No. 5454, pp 782-783.

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