Purification of poly-dA oligonucleotides and mRNA-protein fusions with dT25-OAS resin

Brian Joseph Engel, Brian J. Grindel, Joshua P. Gray, Steven W. Millward

Research output: Contribution to journalArticlepeer-review

1 Scopus citations

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 dT25-conjugated Oligo Affinity Support resin (dT25-OAS) alongside poly-dT14 magnetic beads and dT25-cellulose. dT25-OAS was found to have the highest dA21 oligo binding capacity at 4 pmol/µg, followed by dT14-magnetic beads (1.1 pmol/µg) and dT25-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 dT25-cellulose showed the highest mRNA-affibody purification specificity, followed by dT25-OAS and dT14-magnetic beads. Overall, dT25-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.

Original languageEnglish (US)
Article number126934
JournalBioorganic and Medicinal Chemistry Letters
Volume30
Issue number4
DOIs
StatePublished - Feb 15 2020

Keywords

  • Affibody
  • dT purification
  • mRNA display
  • OAS resin

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Medicine
  • Molecular Biology
  • Pharmaceutical Science
  • Drug Discovery
  • Clinical Biochemistry
  • Organic Chemistry

Fingerprint Dive into the research topics of 'Purification of poly-dA oligonucleotides and mRNA-protein fusions with dT<sub>25</sub>-OAS resin'. Together they form a unique fingerprint.

Cite this