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Enhancers increase transcription efficiency by binding to activator proteins and promoting RNA polymerase activity.
Ref: NCERT Class 12 Biology Chapter 5: Molecular Basis of Inheritance Regulation of Gene Expression - Lac Operon
Discriminating whether two different proteins bind same cis-regulatory element requires resolution of distinct complexes by size. Electrophoretic Mobility Shift Assay allows incubation of radiolabeled probe with each protein individually and in combination; each protein-DNA complex shows unique retarded mobility. Co-occupancy yields ternary complex with further retarded or intermediate mobility compared to binary complexes. Competition with unlabeled DNA and antibody supershift further validates specificity. ELISA and Western blot detect proteins without DNA localization, while ChIP shows in vivo occupancy but lacks in vitro comparative resolution for same-site binding analysis.
Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.
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