Using PAGE to Determine Nucleic Acid Molecular Weight

Molecular weight determination is the most basic use of denaturing polyacrylamide gel electrophoresis. Samples are run versus standards of known molecular weight, and a calibration curve of relative mobility (or distance migrated) versus the logarithm of the size is established. Size can be expressed as molecular weight or number of bases. It is important to bear in mind that sequences of identical length may vary in GC content, and therefore in weight. Additionally, strong base pairing, hairpins and other residual secondary structure may perturb the mobility of a given fragment. Accurate molecular weight determination is best undertaken using size standards which are derived from the same DNA as the sample such as overlapping PCR products, cleavage fragments or synthetic products.

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