Intro
Today, most foods that are sold in grocery stores are Genetically Modified. Genetically Modified Organisms, or GMO's, differ from normal or organic organisms in that they have been specially designed to grow or be a certain way. GMO's are achieved by scientists placing a gene of intrest into the DNA of a particular organism. Farmers have been genetically modifying crops for centuries to encourage specific traits, such as high yield. Genes that are inserted into a plants DNA do not have to even be plant genes at all. Although GM crops seem like a positive alternative, many people would disagree. They say that a super weed or a super insect might be the unintended result, and that weed or insect may be resistant to the pesticides used today. Also, genetically modifying crops may introduce alergens into that food that the buyer might not be aware of. The Technique that we will be using for this Lab involves Polymerase Chain Reaction, or PCR. PCR essentially is a copier for DNA. For instance, a scientist can have trace amounts of a certain fragment of DNA, and after using PCR, they will have millions of copies of that desired strand of DNA. PCR can be used in a variety of situations, from medical diagnosis, to crime scene investigations, and even for aiding sequencing the human genome.
2. Place each tube in a capless microtube adaptor and place in the foam float on ice.
3. Refering to the table and using a fresh tip for each addition, add 20 microliters of the indicated Master Mix to each PCR tube.
4. Refering to the table and using a fresh tip for each tube, add 20 Microliters of the indicated DNA to each PCR tube, be sure to avoid the InstaGene pellet at the bottom of the tubes. Mix by pipetting gently up and down.
5. When instructed, place PCR tubes in thermal cycler.
DAY 3:
1. Set up your gel electrophoresis apparatus as instructed.
2. Obtain your PCR tube from the thermal cycler and place in the capless microtube adaptor. Pulse spin for -3 seconds.
3. Using a fresh tip each time add 10 microliters of Orange G loading dye to each sample and mix well.
4. Load 20 Microliters of the molecular weight ruler and 20 microliters into your gel in the order indicated below:
5. The run time and voltage will depend on the type of gel you are running. Run an agarose gel for 30 mins at 100 V and run a polyachrylamide gel at 200 V for 20 minutes.
6. Stain in Fast Blast DNA stain. Refer to specific instructions depending on gel type.
Procedure
DAY 1:
1. Find your screwcap tubes and label one "Non GMO", and the other "test".
2. Weigh out .5-2g of certified non-GMO food and put it in the mortar and pestle.
3. Add 5 mL of distilled water for every gram of food. The ratio of food to water is achieved from this equation: Mass of Food=____g x 5 = ____ mL
4. Grind with pestle for at least 2 min to form a slurry.
5. Add 5 volumes of water again and mix or grind further with pestle until smooth enough to pipet.
6. Pipet 50 microliters of ground slurry to the screwcap tube containing 500 microliters of InstaGene labeled "non-GMO".
7. Repeat steps 2-5 to prepare the test food sample.
8. Pipet 50 microliters of ground test food slurry to the screwcap tube labeled "test".
9. Shake or flick the non-GMO food and test food InstaGene tubes and place tubes in 95 degree Celsius water bath for 5 minutes.
10. Place tubes in a centrifuge in a balanced conformation and centrifuge for 5 minutes at max speed.
11. Store tubes in a refrigerator until next lesson.
DAY 2:
1. Number PCR tubes 1-6 and initial them. The numbers should correspond to the following tube contents:
DAY 2:
1. Number PCR tubes 1-6 and initial them. The numbers should correspond to the following tube contents:
Tube Number | Master Mix | DNA |
1 (Carrot) | 20 Microliters Plant MM (green) | 20 Microliters Non-GMO food control DNA |
2 (Carrot) | 20 Microliters GMO MM (red) | 20 Microliters food control DNA |
3 (Corn Flour) | 20 Microliters Plant MM (green) | 20 Microliters Test Food DNA |
4 (Corn Flour) | 20 Microliters GMO MM (red) | 20 Microliters Test Food DNA |
5 (Control) | 20 Microliters Plant MM (green) | 20 Microliters GMO Positive Control DNA |
6 (Control) | 20 Microliters GMO MM (red) | 20 Microliters GMO positive control DNA |
2. Place each tube in a capless microtube adaptor and place in the foam float on ice.
3. Refering to the table and using a fresh tip for each addition, add 20 microliters of the indicated Master Mix to each PCR tube.
4. Refering to the table and using a fresh tip for each tube, add 20 Microliters of the indicated DNA to each PCR tube, be sure to avoid the InstaGene pellet at the bottom of the tubes. Mix by pipetting gently up and down.
5. When instructed, place PCR tubes in thermal cycler.
DAY 3:
1. Set up your gel electrophoresis apparatus as instructed.
2. Obtain your PCR tube from the thermal cycler and place in the capless microtube adaptor. Pulse spin for -3 seconds.
3. Using a fresh tip each time add 10 microliters of Orange G loading dye to each sample and mix well.
4. Load 20 Microliters of the molecular weight ruler and 20 microliters into your gel in the order indicated below:
Lane | Sample | Load Volume |
1 | Sample 1: Non-GMO food control with plant primers | 20 Microliters |
2 | Sample 2: Non-GMO food control with GMO primers | 20 Microliters |
3 | Sample 3: Test Food with Plant Primers | 20 Microliters |
4 | Sample 4: Test food with GMO primers | 20 Microliters |
5 | Sample 5: GMO positive DNA with plant primers | 20 Microliters |
6 | Sample 6: GMO positive DNA with GMO primers | 20 Microliters |
7 | PCR Molecular Weight Ruler | 20 Microliters |
8 | Leave Empty | - |
5. The run time and voltage will depend on the type of gel you are running. Run an agarose gel for 30 mins at 100 V and run a polyachrylamide gel at 200 V for 20 minutes.
6. Stain in Fast Blast DNA stain. Refer to specific instructions depending on gel type.
Results
Upon reviewing my own results, it turns out that my test food was a genetically modified food. The corn Flour also, was genetically modified. All in all, this lab was very beneficial and it opened my eyes up to the fact that almost all of the food on the market nowadays is genetically modified.
A+
ReplyDeleteGreat job! Absolutely terrific