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==Conclusion== Because the labeled amount of Enzyme:DNA was calibrated using lambda DNA, which can be considered, for our purposes, randomly assembled, the chances that an enzyme would find the correct 4-mer cutting site is 1/512. This means that the company's enzyme is calibrated for a much lower frequency cutting than for our purposes (which is a guaranteed 1/probe cutting frequency), we have to augment the amount of enzyme used. In this case, for each ug of DNA to be digested we will need roughly 4-5 times the recommended. I used 2x the recommended amount of enzyme, which as the gel showed is not enough. Math note: From a theoretical standpoint, the enzyme is calibrated for a ratio of 1:512 cutting sites:DNA length. Our probes have a ratio of 1:140 which is roughly 3.65x more frequent. Still, given the solution that the enzymes rest in, over-estimating the amount of enzyme needed is a safe move.
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