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J Biotechnol. 2013 Sep 20;167(4):448-53. doi: 10.1016/j.jbiotec.2013.08.006. Epub 2013 Aug 11.

Direct oligonucleotide synthesis onto super-paramagnetic beads.

Author information

  • 1Stanford Genome Technology Center, Stanford University, Palo Alto, CA 94304, USA. Electronic address: m.a.jensen@stanford.edu.

Abstract

Super-paramagnetic beads (SPMB)s used for a variety of molecular diagnostic assays are prepared by attaching pre-synthesized oligonucleotides to the surface via a cumbersome and low efficient method of carbodiimide-mediated amide bond formation. To mainstream the process, we describe a novel procedure of direct oligonucleotide synthesis onto the surface of SPMBs (e.g. MyOne Dynabeads). With the many challenges surrounding containment of paramagnetic beads (≤1 μm) during automated oligonucleotide synthesis, we show that by applying a magnetic force directly to the SPMBs we prevent their loss caused by high-pressure drain steps during synthesis. To date we have synthesized 40 mers using a Spacer 9 phosphoramidite (triethylene glycol) coupled to the surface of hydroxylated SPMBs. HPLC analysis shows successful product generation with an average yield of 200 pmol per sample. Furthermore, because of the versatility of this powerful research tool, we envision its use in any laboratory working with conventional synthesis automation, as employed for single columns and for multi-well titer plates. In addition to direct synthesis of oligodeoxynucleotides (DNA) onto SPMBs, this platform also has the potential for RNA and peptide nucleic acid synthesis.

Copyright © 2013 Elsevier B.V. All rights reserved.

KEYWORDS:

DNA synthesis; Nucleic acid; Super-paramagnetic beads; Synthesis automation

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