Slalom chromatography: size-dependent separation of DNA molecules by a hydrodynamic phenomenon.
Slalom chromatography, a size-dependent DNA fractionation method based on a new principle [Hirabayashi, J., & Kasai, K. (1989) Anal. Biochem. 178, 336-341], was systematically studied in detail. In this method, larger DNA fragments are eluted much later than smaller ones from columns packed with spherical microbeads. Elution of a series of DNA fragments was systematically examined by using columns packed with polymer-based packings of different diameter and different pore size for high-performance gel permeation chromatography. Packings of smaller diameter proved to be superior for resolving the smaller size range of DNA, while the reverse was the case for larger DNAs. Application of a faster flow rate led to larger retardation of every DNA fragment, while at the lowest flow rate applied (0.067 cm/min), all the fragments were eluted almost at the void volume. When the column temperature was lowered, retardation of DNA became larger. On the other hand, differences in the chemical nature and the pore size of packings, or in the hydrophobicity of the eluting solvent, had little effect on DNA retardation. Size-dependent fractionation of DNA was also achieved even on columns packed with nonporous packings having anionic groups (cation exchangers). In conclusion, these results confirmed the previous conclusion that slalom chromatography is not based on an adsorption or equilibrium phenomenon but should be attributed to a hydrodynamic phenomenon.
Hirabayashi
J
J
Department of Biological Chemistry, Faculty of Pharmaceutical Sciences, Teikyo University, Kanagawa, Japan.
Ito
N
N
Noguchi
K
K
Kasai
K
K
D000818:Animals; D010582:Bacteriophage lambda; D002645:Chickens; D002845:Chromatography / Q000379:methods; D003001:Cloning, Molecular; D004247:DNA / Q000302:isolation & purification*; D004279:DNA, Viral / Q000302:isolation & purification; D008099:Liver; D008970:Molecular Weight; D012997:Solvents; D013696:Temperature
D004279:DNA, Viral; D012997:Solvents; D004247:DNA