Purification of Oligonucleotides by Ion-pair Chromatography on Hybrid Silica Particles

Library Number:
Part Number:
Paul Rainville;Martin Gilar;Jeff R Mazzeo;Reb J Russell, II
WCBP 2001; Washington DC; February 20 2001
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Content Subtype:
Trityl-off Oligonucleotides, up to 30-mer
Reaction mixture
XTerra MS C18 2.5 µm Steel 4.6 mm 50 mm
Synthetic oligonucleotides are used as primers for DNA sequencing and PCR and are also being investigated as drug candidates. Due to failure sequences, the purity of 25-mer oligonucleotides is typically 80-85%. Higher purity is required, especially for PCR applications. Typically, oligonucleotides are purified by electrophoresis or HPLC. The current techniques have limitations. Slab-gel electrophoresis is a laborious process, although it affords very high purity (>98%). HPLC suffers from the fact that the oligonucleotides are purified in the trityl on state. After purification, the DMT protecting group must be removed. To address these limitations, we have developed methods for trityl off purification of oligonucleotides using ion-pair reverse phase chromatography on hybrid silica phases (XTerra MSC18). We demonstrate purification strategies for synthetic oligonucleotides up to 30-mers on a micromolar scale.

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