Analysis and purification of synthetic large oligodeoxyribonucleotides by HPLC on RPC-5 like resin

Large oligodeoxyribonucleotides (20-160 bases), synthesized by the phosphoramidite method, have been analyzed and purified by HPLC on a RPC-5 like resin (Neosorb LC). Linear gradient of NaClO4 solution containing 10 mM NaOH and 0.1 mM EDTA was carried out for the elution. Large oligodeoxyribonucleotides bearing 4,4'-dimethoxytrytyl (DMT) group were separated very well from the shorter failed by-products. After removal of the DMT group, the products were analyzed and purified by repeating HPLC on Neosorb LC. This HPLC system gave well resolution of the desired oligodeoxyribonucleotide (over 50 bases) from the base modified by-products with the same chain length. The chromatogram showed the presence of large amount of by-products in addition to the desired product when methylphosphoramidite method was employed for the DNA synthesis.