Gelshift Chemiluminescent EMSA

The Gelshift Chemiluminescent EMSA Assay Kit provides a simple, non-radioactive assay to identify protein-DNA binding with proven reagents. In this electrophoretic mobility shift assay (EMSA), cell extracts or purified factors are incubated with biotin end-labeled probe containing the consensus binding site of interest. Samples in which the protein of interest bound the target DNA will migrate slower than DNA alone resulting in a "shift" of the labeled DNA band.

Gelshift Chemiluminescent EMSA Method

The Gelshift Chemiluminescent EMSA Kit provides a non-radioactive method to detect DNA-protein interactions. In this method, cell extracts or purified factor are incubated with a biotin 3' or 5' end-labeled DNA probe containing the consensus binding site of interest. Samples are then resolved by electrophoresis on a native polyacrylamide gel and transferred to a nylon membrane. The biotin end-labeled DNA probe is detected using streptavidin conjugated to horseradish peroxidase (HRP) and a chemiluminescent substrate. Samples in which the protein of interest bound the target DNA will migrate slower than DNA alone resulting in a "shift" of the labeled DNA band.

Example EMSA Binding Reactions:

1.Biotin-labeled DNA = no shift in the absence of cell extract or purified factor

2.Biotin-labeled DNA + Extract = DNA shift due to the binding of protein to the labled DNA probe

3.Biotin-labeled DNA + Extract + Excess Unlabeled DNA = no shift as the excess of unlabeled DNA competes for binding of the target protein in the extract; this reaction verifies the specificity of the protein-DNA interaction

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Gelshift Chemiluminescent EMSA