WET LAB DNA Barcoding: From Samples to Sequences - Northwest ...

WET LAB
KEY
• Laboratory math
• Analyzing DNA gel results, including estimating DNA concentration
• Diluting samples
7. How is this PCR purification similar to the DNA purification that you performed? How is it different?
The ultimate goal of DNA purification and PCR purification is the same: to obtain purified DNA for future
experiments. In both cases, students are separating DNA from other contaminants or components using a spin
column (i.e., DNA-binding membrane). As in DNA purification, in PCR purification, samples are loaded onto a
DNA-binding spin column, washed, and then eluted (removed) from the column for future use.
These two experimental techniques have key differences. In DNA purification, there are additional steps
involved compared to PCR purification, including lysis of the cell, removal of cell debris, and two different
wash steps (a pre-wash and a wash). In the PCR purification, there is no cell to lyse. In DNA purification, the
contaminants, or things being removed from the DNA, include cellular organelles and membranes. In PCR
purification, the contaminants, or things being removed from the DNA, are the components of the PCR (Taq
DNA polymerase, unused primers, unused dNTPs, and buffer). Students may also note that the DNA recovered
in the DNA purification is total cellular DNA (i.e., nuclear and mitochondrial DNA), and the DNA is eluted in
Elution Buffer, while the DNA recovered in the PCR purification is the amplified (or copied) PCR product, and
the DNA is eluted in pure water.
Using Bioinformatics: Genetic Testing
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©Northwest Association for Biomedical Research—Updated October 2012