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PCR with modifications
This week we ran multiple gels to see if we could get banding of our DNA. In both gels we ran no banding was found. We ran with higher concentrations of DNA, longer extension times, and lower PCR temperatures. We also attempted to make a digestion of our plasmid DNA for future classes to use.
This is the gel of the PCR that we had ran 5 microliters of DNA sample #3, and lowered our PCR temperatures 5 degrees overall. No bands were seen in the in the sample lanes except for A3. Starting from the left lane: Positive control, Negative control, A3 sample, 1000 Bp ladder, A2 sample, and A1 sample. A1 was at 60 degrees, A2 was at 50 degrees, A3 along with the Neg. and Pos. controls were at 45 degrees. The Positive control was seen at the 300 Bp mark.
This is the gel of the PCR where we used 8 microliters of DNA instead of 5 microliters. We had to use our samples #1 and #2 combined because we ran out of sample #3. We also made the extension time in the PCR for 5 minutes. to amplify our gene again. There were no bands appearing except for the positive control. Starting from the left to right: the first lane is the positive control (band seen at 300 Bp mark), negative control, A5, A4, 1000 Bp ladder, A3, A2, A1. The A1 sample was in the PCR at 65 degrees C, A2 at 61 degrees, A3 at 57 degrees, A4 at 48.8 degrees, and A5 and the controls were at 45 degrees.