BC002 – TRIM31 PCR Optimization

Submitted by Brittnee Crane

A PCR for HA-pcDNA3.1(+)-TRIM31 was made up using the Q5 High Fidelity Master Mix protocol, and there were some adjustments made to the thermocycler’s settings. We changed the annealing temperature and the denaturation time, in hopes that we would obtain the properly sized band would appear when ran on the gel. The PCR product was then run on a gel and no promising results were obtained. A PCR was made for HA-pcDNA3.1(+)-TRIM69 and Dr. Graff ran it on a gel for a longer period of time than in the previous experiment, hoping that the two bands within the “long” band would be spaced further apart. The longer run time was successful in separating the two bands. The band displaying at ~1.5Kb was cut out of the gel and put through the gel purification process. The gel purified product was run on a gel along with a digest of pGADT7-empty, and the concentrations of both were estimated using crazy units.

BC002

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