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Electrophoretic mobility of λ phage HIND III and HAE III DNA fragments in agarose gels: A detailed study
Author(s) -
Hervet H.,
Bean C. P.
Publication year - 1987
Publication title -
biopolymers
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.556
H-Index - 125
eISSN - 1097-0282
pISSN - 0006-3525
DOI - 10.1002/bip.360260512
Subject(s) - electrophoresis , agarose , chemistry , ionic strength , ionic bonding , reptation , electric field , dna , gel electrophoresis of nucleic acids , field strength , base pair , persistence length , chemical physics , analytical chemistry (journal) , chromatography , molecule , polymer , aqueous solution , biochemistry , physics , ion , organic chemistry , quantum mechanics , magnetic field
The electrophoretic mobility (μ) of DNA fragments from λ phage and ΦX 174, split by restriction enzyme to molecular lengths from 3 × 10 2 to 2.36 × 10 4 base pairs, has been investigated in 0.6–4% agarose gels at various field strengths, ionic strengths, and temperatures. As already observed, μ is seen to be very sensitive to the field, increasing with field strength. The sensitivity increases with the molecular length of the DNA and decreases at high gel concentration. Our data are in qualitative agreement with recent theoretical predictions that concern the influence of the electric field on electrophoretic mobility. Mobility data have been extrapolated to zero field. This enables a comparison of our experimental results with theoretical predictions on the dependence of μ on the molecular weight of the DNA fragments. Our data fit, quite closely, a reptation model, where the tube path is described as a semiflexible entity with a persistence length equal to the pore diameter. The influence of the agarose concentration and the ionic strength of the buffer on the two parameters of the model—intrinsic electrophoretic mobility (μ 0 ) and the number of base pairs per element of the tube ( g )—are well described by the model. The temperature dependence of the electrophoretic mobility, together with the influence of the agarose concentration on μ 0 , indicate that the hydrodynamic drag is the leading frictional force on the DNA molecules in the gel.