What is the result of using only SDS (no reducing agent) in PAGE?
Accurate molecular weight estimation by SDS-PAGE requires both anionic detergent and reducing thiol for complete linearization. Sodium dodecyl sulfate disrupts non-covalent interactions and imparts uniform negative charge but cannot cleave covalent disulfide linkages between cysteine residues, which constrain polypeptide loops and maintain compact domains. Addition of beta-mercaptoethanol or dithiothreitol reduces disulfides to free sulfhydryls, fully extending chain. Without reducing agent, proteins remain denatured yet partially constrained with intact disulfides, resulting in anomalously fa
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.