Abstract
Solid-phase resins functionalized with poly-deoxythymidine (dT) oligos facilitate purification of poly-adenylated molecules from solution through high affinity, high selectivity base-pairing interactions. These resins are commonly used to purify messenger RNA (mRNA) from complex biological mixtures as well as mRNA-protein fusion molecules for mRNA Display selections. Historically, dT-conjugated cellulose was the primary resin for poly-dA purification, but its scarcity has prompted the development of alternative resins, most notably dT-functionalized magnetic beads. In order to develop a cost-effective alternative to commercially available poly-dT resins for large-scale purifications of mRNA-protein fusions, we investigated the purification properties of dT(25)-conjugated Oligo Affinity Support resin (dT(25)-OAS) alongside poly-dT(14) magnetic beads and dT(25)-cellulose. dT(25)-OAS was found to have the highest dA(21) oligo binding capacity at 4 pmol/µg, followed by dT(14)-magnetic beads (1.1 pmol/µg) and dT(25)-cellulose (0.7 pmol/µg). To determine the resin specificity in the context of a complex biological mixture, we translated mRNA-protein fusions consisting of a radiolabeled Her2 affibody fused to its encoding mRNA. Commercial dT(25)-cellulose showed the highest mRNA-affibody purification specificity, followed by dT(25)-OAS and dT(14)-magnetic beads. Overall, dT(25)-OAS showed exceptionally high binding capacity and low background binding, making it an attractive alternative for large-scale mRNA purification and mRNA Display library enrichment.