Cloning TRIM31 and TRIM69 into pGADT7

Submitted by Brittnee Crane

Summary: PCR reaction mixtures were made up for pcDNA3.1(+)-HA-TRIM31 variant 1 and pCAGGS-HA-TRIM69 and put through the thermocycler with the above settings. Dr. Graff ran the two PCR samples on a 1% agarose gel and TRIM31 showed as primer dimers and TRIM69 had two different bands appear, one short and one long. Both bands were cut out and put through the gel purification with a Qiagen kit. It is hard to see in the images, but the long band from TRIM69 contained two bands.


Dr. Graff prepared a digest of pGADT7 cut with EcoR1 and BamH1. The digest and the gel purified TRIM69 samples were then run on a gel together. The concentration of the digest was estimated from the gel, but we decided to repeat the TRIM69 PCR, and run in again in hopes that the two bands within the “long band” would separate given a longer running time on the gel.


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