Stretching and overstretching of DNA in pulsed field gel electrophoresis. I. A quantitative study from the steady state birefringence decay

Abstract

SYNOPSISUsing a sensitive birefringence instrument, the birefringence arising from the orientation of the DNA chain during electrophoretic transport has been recorded. This birefringence is shown to proceed both from the alignment (stretching) of the molecule in the direction of the electric field and from the extension of the length of its primitive path (overstretching) . The contribution of these two processes can be separated in the decay of the birefringence after the end of the application of the electric field. The fast relaxation of the overstretching occurs first and is demonstrated to be the main contribution to the birefringence. The orientation factor of the remaining stretched state and its decay can be quantitatively understood using the biased reptation model. It provides, in addition, a high value for the tube diameter or gel pore size a (4500 k 450 A for a 0.7% agarose gel with a c;'.~ dependence in the agarose concentration cg) and a low value for the effective charge per base pair (0.2e as compared to 0.5e using the condensation hypothesis). The contribution of overstretching to the birefringence is also quantitatively interpreted in term of the change in the mean length 1 of DNA inside a pore size a. The dynamics of decay of this overstretching is well represented by a stretched exponential with a stretching exponent a = 0.44. The mean decay time decreases slightly with increasing fields and scales with the overall DNA length close to N ; . 0 1993 John Wiley & Sons, Inc.

Cyclic migration of DNA in gels: DNA stretching and electrophoretic mobility

“…The study by Weill and co-workers [21] has shown that the field strength and DNA size dependence of the tube orientation is in good agreement with the reptation theory, and Stube can therefore be expected to follow the predicted decrease as the pores become smaller in denser gels. This proposal has not been tested directly by measurements of Stub, at different gel concentrations, and may seem to be contradicted by the observation that the total DNA orientation, S , goes through a maximum…”

“…3) [20,21,261. In this kind of experiment the molecules are first allowed to reach equilibrium in the gel by turning off the field for tens of minutes [26].…”

Migration of single‐stranded DNA in polyacrylamide gels during electrophoresis

“…(4), i.e., For that purpose, one defines the ratio (33) where F(a/l) = 1-3 a/NLang-'(all) where Lang-' is the inverse Langevin function. [12] for dsDNA in agarose gels [13]. The time z', is an important dynamical parameter when devising pulse trains for pulsed field gel electrophoresis.…”

“…Following the procedure described by Mayer et al [12], the contribution of overstretching to birefringence, as taken from the second term in the bracket of Eq. (4), i.e., For that purpose, one defines the ratio (33) where F(a/l) = 1-3 a/NLang-'(all) where Lang-' is the inverse Langevin function.…”

Influence of optical probing with YOYO on the electrophoretic behavior of the DNA molecule

“…

Influence of optical probing with YOYO on the electrophoretic behavior of the DNA molecule

Department ofThe influence of the fluorescent dye YOYO (1,[1][2][3][4]4,8,8,-tetramethylChahers University of

4,8-diazaundecamethylene)bis[4-[[3-methyl-benzo-1,3-oxazol-2-yl]methylidene]-Goteborg, Sweden 1,4-dihydroquinolinium] tetraiodide) on the electrophoretic behavior of the DNA molecule was investigated. This is important when using YOYO as a probe in capillary electrophoresis or in fluorescence microscopy studies of DNA with the purpose of studying the migration mechanism of DNA on the molecular level.

…”