Pulsed-field polyacrylamide gel electrophoresis: basic phenomena and applications.
Pulsed-field gel electrophoresis (PFGE) using polyacrylamide gels, termed pulsed-field polyacrylamide gel electrophoresis (PF-PAGE), had been developed for the effective separation of linear DNAs from circular ones [1]. The first generation PF-PAGE employed horizontal polyacrylamide gels run in a contour-clamped homogeneous electric field (CHEF) apparatus. The second generation system, using a vertical slab gel in a discontinuous buffer system and field inversion gel electrophoresis (FIGE), was found to be easier to handle and requires a much shorter time for separation than the previous one [2]. In this report, basic aspects of the second generation PF-PAGE, such as the effects of a discontinuous buffer system and field inversion on the DNA migration in polyacrylamide gels, were investigated. The results indicate that the periodic inversion of electric field can broaden the resolving capability of polyacrylamide gels, enabling DNAs that otherwise fail to enter polyacrylamide gels to be resolved in such systems. Successful and possible applications of PF-PAGE techniques are also discussed.
Ito
T
T
Laboratory of Molecular Medicine, University of Tokyo, Japan.
Hohjoh
H
H
Sakaki
Y
Y
D001483:Base Sequence; D002021:Buffers; D003001:Cloning, Molecular; D004247:DNA / Q000302:isolation & purification*; D004270:DNA, Circular / Q000302:isolation & purification*; D004560:Electricity; D016521:Electrophoresis, Gel, Pulsed-Field / Q000379:methods*; D006801:Humans; D008969:Molecular Sequence Data; D012967:Sodium Dodecyl Sulfate
D002021:Buffers; D004270:DNA, Circular; D012967:Sodium Dodecyl Sulfate; D004247:DNA