Separation and chain-length determination of polynucleotides by gel filtration

Gel filtration provides an easy method for chain-length determination in homologous series of oligonucleotides defined by base composition and degree of phosphorylation under various elution conditions.

Abstract

Homologous series of oligonucleotides, defined by base composition and degree of phosphorylation, were chromatographed on Sephadex G-25, G-50, and G-75 under various elution conditions. The application range of each gel can be modified extensively by varying the ionic strength of the elution buffer. Kd. values of polynucleotides decrease logarithmically with increasing chain-length. Kd values are determined mainly by size and net charge of the molecules (proportional to chain-length) and less by base-specific interactions (not proportional to chain-length). Short-chain polymers can be separated according to chain-length, base composition and degree of terminal phosphorylation. Higher polynucleotides are separated mainly according to chain-length. Therefore in this range gel filtration provides an easy method for chain-length determination.