Once genomic DNA is extracted, PCR can be performed to amplify a specific region of the DNA. PCR reactions can be set up by preparing a master mix of reactants to be added to the DNA. This master mix includes sterile water, polymerase buffer, a dNTP nucleotide mix, two primers for either end of the selected fragment, and Taq polymerase. The Taq polymerase is always added to the mix last and is treated with care to prevent denaturation of the protein. The master mix and DNA are then added to PCR tubes, which are small and typically only hold about 2 milliliters of liquid at maximum. The tubes are placed in a PCR machine that will cycle through a set protocol of specific temperatures at specific times. Different temperatures are required to repeatedly denature the double stranded DNA, anneal the primers to the DNA, and extend the DNA with the polymerase. Once PCR is complete, which usually takes 2-3 hours, the PCR products can be run on a gel to verify the fragment and separate the DNA from the contaminants in the PCR reaction. Agarose gel electrophoresis is completed, loading the gel wells with a DNA ladder and each sample mixed with loading dye. Once the gel has been run, the DNA can be visualized using UV light. There should be a distinct band that represents your DNA.
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