Separation of nucleic acids by high-performance ion-exchange chromatography.

ABSTRACT

Separation of various nucleic acids was evaluated by high-performance ion-exchange chromatography on non-porous resin, TSKgel DNA-NPR. A 1 kb ladder DNA was studied as a model DNA on operational variables like flow rate, gradient time, temperature, sample load, etc.. As results, various DNA fragments were well separated within 15 min by 20 min linear gradient at a flow rate between 0.5 and 0.75 ml/min at room temperature while the resolutin was dependent on molecular weight of the sample. The relationship between sample load and its peak area was examined on polymerase chain reaction (PCR) product. The product was found to be quantitatively recovered even with nanogram loads. The detection limit was 3.8 ng at signal to noise level (S/N) of 3. This non-porous ion-exchanger also showed high resolution on separation of ther nucleic acids like transfer RNA, oligonucleotides (single-stranded) DNA.