How does the assay work?
Biotin end-labeled DNA containing a putative or known binding site is incubated with a nuclear extract or purified factor. This reaction is then subjected to gel electrophoresis on a native polyacrylamide gel and transferred to a nylon membrane. Because DNA-protein complexes migrate slower than DNA alone in a native gel, a “shift” in the migration of the labeled DNA occurs. The biotin end-labeled DNA is detected using a streptavidin-horseradish peroxidase conjugate and a chemiluminescent substrate developed for the LightShift Kit. The signal is then detected with X-ray film or a high quality CCD camera.
