Part 3a: Genetically Modified Soy
Identification of Food Product
Materials & Equipment
Thermocycler
Mini benchtop centrifuge
Vortexer
Pipets – 2 µL, 20 µL, 200 µL, 1000 µL + tips
Freezer Blocks (1 per group)
PCR Tubes
PCR Product
Sterile Water
Endogenous Probe
Transgenic Probe
0.25 M NaCl
Overview: We are using two different types of
probes to determine whether the food product is
endogenous (natural) or transgenic (genetically
modified).
Time: 45 minutes for both a and b parts
Protocol
1. Prepare two PCR tubes for each of your group’s soy samples by labelling one as “N” for Non GMO and
one as “G” for Genetically Modified. We will test each sample for the presence of the naturally occuring
and the genetically modified gene.
2. Fill the appropriate tubes with the reagents mentioned in the tables below.
Preparation of “N” Tube
Reagent
PCR Product
0.25 M NaCl
Poly-A Tailed Probe
Sterile H20
Volume ✔
5 µL
4 µL
1 µL
10 µL
Preparation of “G” Tube
Reagent
PCR Product
0.25 M NaCl
Poly-A Tailed Probe
Sterile H20
Volume ✔
5 µL
4 µL
1 µL
10 µL
3. Vortex (approximately 10 seconds) and centrifuge each tube for 1-2 seconds.
4. Place all tubes in the thermocycler.
5. Run the thermocycler using the following reaction conditions:
Temperature (°C)
95
ICE
25
Time
5 minutes
1 minute
10 minutes
NOTES: -Record in your worksheet the location of your samples in the thermocycler for future reference.
-When moving tubes from the ice back to the thermocycler, make sure that the heating compartment
is already at 25 ⁰C. In some cases this may take longer than 1 minute.
6. Follow “Biosensor Setup” on following page while tubes are in the thermocycler.
Protocol was developed by undergraduates, graduates and post docs at Tufts University with funding from David R. Walt’s
Howard Hughes Medical Institute Professor’s Award.
Part 3b: Genetically Modified Soy
Biosensor Readout
Materials
Pipets – 10 µL, 20 µL, 1000 µL
Sonicator
Completed Biosensors
Running Buffer*
Beads
Thermocycler
“N” and “G” Tubes
Microcentrifuge Tubes
Overview: The probes we added to the sample
DNA will be detected using the biosensor. Capillary
action brings the solution up the biosensor then the
probes react with the line on the biosensor to
produce the result.
*See GMO buffer sheet for buffer recipes.
Biosensor Setup
1. Obtain one biosensor per tube.
2. Line up the biosensors on a clean surface.
3. Prepare a 1.5 mL microcentrifuge tube for each biosensor by cutting off the caps and labeling each
tube with “N” or “G”. (You may also choose to label the top of the biosensor with this information)
4. Add 300 µL of Running Buffer to each tube.
5. Separate beads into 24 µL aliquots. This is enough for four biosensors.
NOTE: Only 20 µL should be used for four biosensors, but beads are inevitably lost during the sonication and
pipetting processes.
Running the Biosensor
1. The aliquot of beads (24 µL) must be sonicated for 10 seconds right before they are spotted onto the
biosensor.
2. Working quickly, add 10 µL of the GM Plasmid Detection
mixture to the Lateral Flow Biosensor at the top of the
glass fiber.
3. Add 5 µL of beads just above the glass fiber portion of the
biosensor.
NOTE: Add the solutions slowly to prevent them from traveling down to
the lower cellulose pad and getting stuck. It is alright if the solutions
start to travel up the membrane before the biosensor is placed in the
Running Buffer.
4. Place the biosensor in the corresponding 1.5 mL tube.
5. Signal will appear 15-20 minutes after biosensor has been
placed in Running Buffer
solution.
Protocol was developed by undergraduates, graduates and post docs at Tufts University with funding from David R. Walt’s
Howard Hughes Medical Institute Professor’s Award.