UNIT 10.14 Purification of Synthetic Oligonucleotides via Catching by Polymerization
Published Online: 1 JUN 2012
Copyright © 2012 by John Wiley & Sons, Inc.
Lab Protocol Title
Current Protocols in Nucleic Acid Chemistry
How to Cite
Fang, S., Fueangfung, S. and Yuan, Y. 2012. Purification of Synthetic Oligonucleotides via Catching by Polymerization. Current Protocols in Nucleic Acid Chemistry. 49:10.14.1–10.14.21.
- Published Online: 1 JUN 2012
This unit describes the purification of synthetic oligodeoxyribonucleotides (ODN) using a catching-by-polymerization approach. In a crude ODN, the major impurity is the failure sequences generated in the coupling step of each synthetic cycle. They are difficult to remove due to the similarity of their physical properties to the full-length sequences. Two non-chromatographic methods are described in the unit to solve the problem. In the first one, during automated synthesis, the failure sequences are tagged with a methacrylamide group, which is polymerizable and can participate in acrylamide radical polymerization reactions; the full-length sequences are not tagged. After synthesis, the crude mixture is subjected to polymerization. The failure sequences are incorporated into an insoluble polymer; the full-length sequences are extracted with water. In the second method, the full-length sequences are tagged with a methacrylamide group via a cleavable linker; the failure sequences are not tagged. After synthesis, the full-length sequences are incorporated into a polymer; the failure sequences are washed away with water. Pure full-length sequences are cleaved from the polymer. The two methods are complementary. Curr. Protoc. Nucleic Acid Chem. 49:10.14.1-10.14.21. © 2012 by John Wiley & Sons, Inc.
- catching by polymerization;
- scalable purification;
- acrylamide radical polymerization;
- full-length sequence;
- failure sequence