Unit

UNIT 3.17 Release of DNA Oligonucleotides and Their Conjugates from Controlled-Pore Glass Under Thermolytic Conditions

  1. Andrzej Grajkowski1,
  2. Jacek Cieślak1,
  3. Scott Norris2,
  4. Darón I. Freedberg2,
  5. Jon S. Kauffman3,
  6. Robert J. Duff3,
  7. Serge L. Beaucage1

Published Online: 1 DEC 2008

DOI: 10.1002/0471142700.nc0317s35

Current Protocols in Nucleic Acid Chemistry

Current Protocols in Nucleic Acid Chemistry

How to Cite

Grajkowski, A., Cieślak, J., Norris, S., Freedberg, D. I., Kauffman, J. S., Duff, R. J. and Beaucage, S. L. 2008. Release of DNA Oligonucleotides and Their Conjugates from Controlled-Pore Glass Under Thermolytic Conditions. Current Protocols in Nucleic Acid Chemistry. 35:3.17:3.17.1–3.17.21.

Author Information

  1. 1

    Division of Therapeutic Proteins, Center for Drug Evaluation and Research, Food and Drug Administration, Bethesda, Maryland

  2. 2

    Division of Bacterial, Parasitic & Allergenic Products, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland

  3. 3

    Lancaster Laboratories, Lancaster, Pennsylvania

Publication History

  1. Published Online: 1 DEC 2008
  2. Published Print: DEC 2008

Abstract

The sequential functionalization of long-chain alkylamine controlled-pore glass (CPG) with a 3-hydroxypropyl-(2-cyanoethyl)thiophosphoryl linker and a dinucleoside phosphorotetrazolide leads to a uniquely engineered support for solid-phase synthesis. Unlike conventional succinylated-CPG supports, this support is designed to allow oligonucleotide deprotection and elimination of deprotection side-products to proceed without release of the oligonucleotide. When needed, the DNA oligonucleotide can be thermolytically released in 2 hr under essentially neutral conditions. The modified CPG support has been successfully employed in the synthesis of both native and fully phosphorothioated DNA 20-mers. On the basis of reversed-phase HPLC and electrophoretic analyses, the purity of the released oligonucleotides is comparable to that of identical oligonucleotides synthesized from succinylated-CPG supports, in terms of both shorter-than-full-length oligonucleotide contaminants and overall yields. The detailed preparation of DNA oligonucleotides conjugated with exemplary reporter or functional groups, either at the 3′-terminus or at both 3′- and 5′-termini, is also described. Curr. Protoc. Nucleic Acid Chem. 35:3.17.1-3.17.21. © 2008 by John Wiley & Sons, Inc.

Keywords:

  • deoxyribonucleoside phosphorodiamidites;
  • solid-phase synthesis;
  • modified CPG support;
  • thermolytic conditions;
  • DNA oligonucleotides conjugates