FAQ: If the DNA resulting from a Topoisomerase I reaction is run in an ethidium bromide gel and there seems to be no evidence of enzyme activity, what is the most probable explanation?

Ethidium bromide intercalates between the bases of the DNA and induces supercoiling. All relaxed plasmids regardless of linking number will become supercoiled in the presence of ethidium bromide and appear to run at the same position equivalent to any supercoiled DNA in the sample. Therefore to check the result of a Topoisomerase I reaction, the products MUST be run in a gel without any ethidium bromide. After the gel is run, it can then be stained with ethidium bromide to visualize the DNA. The relaxed forms of the plasmid will run slower and at the top of the gel. Any supercoiled plasmid will run faster and lower on the gel.