A model for the mobility of single-stranded DNA in capillary gel electrophoresis.
The electrophoretic mobility of single-stranded DNA in denaturing polyacrylamide gel-filled capillaries is analyzed as a function of the applied electric field. The resultant mobility plots are complex functions of the fragment size and electric field. Traditionally, these plots are separated into three mobility regimens corresponding to Ogston sieving, reptation without stretching and reptation with stretching theories. However, none of these theories accurately models the variations in mobility that we observe with electric field strength. As a result, we propose a modification of the Ogston sieving theory which accounts for the stretching of migrating DNA molecules in the direction of the electric field. This theory assumes that the applied electric field in conjunction with the gel matrix distorts the DNA, altering the effective size of the migrating molecule. The stretched DNA offers a smaller cross-section to the gel pore and thus sieves as though it were a smaller molecule. In this modified Ogston theory, the electrophoretic mobility depends only on the applied electric field, the size of the fragment, and constants which are independent of size and field strength. The modified Ogston equation accurately predicts the mobilities of DNA fragments in all three mobility regimes, providing a single, simple model to account for all of the observed behavior.
Luckey
J A
JA
Department of Chemistry, University of Wisconsin 53706.
Smith
L M
LM
D004277:DNA, Single-Stranded / Q000737:chemistry*; D004574:Electromagnetic Fields; D004586:Electrophoresis / Q000379:methods; D006358:Hot Temperature; D008956:Models, Chemical
D004277:DNA, Single-Stranded