Evolved polymerases facilitate selection of fully 2′-OMe-modified aptamers†

Abstract

Evolved DNA polymerases are used in selections with fully 2′-OMe modified libraries to identify aptamers with high affinity for HNE.

Abstract

RNA or DNA aptamers with 2′-OMe-modifications have been pursued to increase resistance to nucleases, but have been difficult to identify because the OMe groups ablate polymerase recognition. We recently reported evolution of the thermostable DNA polymerases SFM4-6 and SFM4-9, which enable the efficient “transcription” and “reverse transcription”, respectively, of 2′-OMe oligonucleotides. With these polymerases, we now report the first selection of fully 2′-OMe modified aptamers, specifically aptamers that bind human neutrophil elastase (HNE). Two aptamers, 2mHNE-1 and 2mHNE-2, were isolated after five rounds of selection, and four more, 2mHNE-3–6, after an additional five rounds that included selection pressure for binding in the presence of serum. All six aptamers bind with reasonable affinity, which requires the 2′-OMe substituents. Further characterization of one aptamer, 2mHNE-5, showed that unlike a previously reported natural anti-HNE aptamer, affinity for HNE is retained in the presence of high concentrations of salt or serum. The polymerases SFM4-6 and SFM4-9 should prove valuable for the production and further exploration of modified aptamers.

Notes

†Electronic supplementary information (ESI) available: Methods, supplementary scheme, tables and figures. See DOI: 10.1039/c7sc03747c