Rationale

Today we will run gel electrophoresis on the PCR samples created on 10/31/18.

Procedure

• Established an aseptic zone.
• 40 mL of 1X TBE and 0.8 g of Agarose were added into a flask and microwaved until clear.
• The mixture was set aside to cool for 5 minutes. After cooling, 2 µL of EtBr was added to the flask and was swirled around.
• The contents in the flask were poured into a gel apparatus with a comb inserted and set aside for 20 minutes.
• After the gel had set, the rubber sides of the apparatus were removed as well as the comb. 1X TBE was used to submerge the gel.
• 2.5 µL of dye was added to each PCR tube before pipetting into the wells.
• 10 µL of microcentrifuge tubes 1 •, 2 •, 3 •, positive control 1, positive control 2, and positive control 3 were pipetted into wells 1, 2, 3, 5, 6, and 7 respectively.
• 5 µL of DNA ladder was pipetted into well 4.
• The gel electrophoresis apparatus was plugged into a power source, which was then turned on and set aside for 30 minutes.
• The gels were then imaged to reveal the results.

Observations

After the gel was plugged into the power source, the blue color traveled approximately the same distance across the gel for each sample and positive control.

Conclusions/Next Steps

The results were negative, however, one of the “phage DNA” samples tested was a known positive. This indicates that the gel electrophoresis method for testing for phage presence is flawed.