High-resolution liquid chromatography of DNA fragments on non-porous poly(styrene-divinylbenzene) particles

Abstract

DNA restriction fragments and PCR products were separated by means of ion-pair reversed-phase high-performance liquid chromatography on alkylated non-porous poly(styrene-divinylbenzene) particles with a mean diameter of 2.1 μm. Optimum resolution was obtained by using an acetonltrile gradient in 100 mM of triethylammonlum acetate and a column temperature of 50 °C. This allowed the separation of DNA fragments differing in chain length by 1 - 5% up to a size of 500 base pairs. PCR products could be analyzed directly in less than two minutes with a concentration sensitivity of at least 300 ng/ml. Compared with anlon-exchange chromatography or gel electrophoresls no desaltation of the purified DNA molecules is required because the volatile buffer system can be readily evaporated. Subsequently, the method was used for the semiquantltative evaluation of the expression of multidrug resistance genes in mononuctear white blood cells. © 1993 Oxford University Press.