Which electrophoresis is best for detecting isomers of same protein?
Proteins encoded by single gene often exist as multiple isoforms created by alternative splicing, phosphorylation, acetylation, deamidation or limited proteolysis, producing subtle variations in charge and mass indistinguishable in one-dimensional systems. SDS-PAGE masks charge differences and poorly resolves small mass changes, while isoelectric focusing alone cannot separate mass variants. Two-dimensional electrophoresis combining isoelectric focusing for pI separation and SDS-PAGE for molecular mass provides orthogonal resolving power, displaying isoforms as distinct spots across pH and siz
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.