A rapid and effective method for separating oligonucleotides using high-performance ion-exchange chromatography is described. Columns were prepared by adsorbing a layer of polyethylene imine on 5-micron-diameter porous silica followed by crosslinking with a multifunctional oxirane. This weak anion-exchange matrix was found to be useful in the separation of mononucleotides and oligonucleosides containing up to 20 residues in either homo- or heteropolymers. Small analytical columns (4.2 X 150 mm) had a capacity ranging from less than a microgram in analytical separations to several milligrams in the preparative mode. The columns have proven to be especially useful in assessing the purity of precursor blocks, monitoring the chemical synthesis of oligonucleotides, and isolating reaction products after the synthesis.
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