CGDP - DNA Amplification
Part 2

Extraction Amplification Sequencing

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FLMNH 2005


 
  DNA amplification takes place in small tubes placed into a PCR machine like the one to the left. It is basically a fancy heating and cooling block. The mix in the tubes initially contains a small amount of Genomic DNA, Taq polymersae, primers, salts, buffers, and extra G's, A's, T's and C's that are added on while the DNA pieces are built. Once the cycles finish we visualize the reactions to see if they worked by running a small amount out on an agarose gel.
   The image to the right shows some of these reactions run out on a gel and stained with ethidium bromide which attaches to DNA and fluoresces under UV light. Some reactions work better than others and because there are more copies, these reactions fluoresce more strongly. Some reactions don't work at all, and you try again. A blank, or neagtive control, is also cycled to be certain that there is no contamination. Additionally a standard 100 base pair ladder is run in a lane to measure the lengths of the amplified piece.
   Once the original PCR reactions are visualized, then it is time to move on to cleaning up the reactions and sequencing the target gene.