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Updated: Jul 5, 2026

Nucleoside Triphosphates - From Synthesis to Biochemical Characterization

Nucleoside Triphosphates - From Synthesis to Biochemical Characterization

Published on: April 3, 2014

Nucleoside phosphoramidites containing cleavable linkers.

Richard T Pon¹

¹University of Calgary, Calgary, Alberta, Canada.

Current Protocols in Nucleic Acid Chemistry

|April 23, 2008

Summary

New phosphoramidite reagents enable efficient oligonucleotide synthesis on inexpensive supports. This method simplifies DNA synthesis, producing desired products like PCR primers and double-stranded DNA in a single operation.

Area of Science:

Oligonucleotide Synthesis
Chemical Biology
Biotechnology

Background:

Traditional oligonucleotide synthesis often requires specialized supports and additional dephosphorylation steps.
The use of phosphoramidite reagents is standard, but limitations exist in support compatibility and product diversity.
Developing cost-effective and versatile methods for oligonucleotide synthesis remains a key challenge.

Purpose of the Study:

To introduce novel phosphoramidite reagents for simplified oligonucleotide synthesis.
To enable the use of inexpensive, underivatized solid supports for universal oligonucleotide production.
To develop a method for producing multiple linked oligonucleotides, including PCR primers and double-stranded DNA, efficiently.

Main Methods:

Utilized phosphoramidite reagents with a cleavable 3'-ester linkage for attaching the first nucleoside to solid supports.

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Last Updated: Jul 5, 2026

Nucleoside Triphosphates - From Synthesis to Biochemical Characterization

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Chemical Triphosphorylation of Oligonucleotides

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Protocol for the Solid-phase Synthesis of Oligomers of RNA Containing a 2'-O-thiophenylmethyl Modification and Characterization via Circular Dichroism

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Published on: July 28, 2017

Employed inexpensive, underivatized supports like Long Chain Aminoalkyl Controlled Pore Glass (LCAA-CPG).

Developed specific phosphoramidites with succinate and sulfonyldiethanol linkages for creating 3'-OH and 5'-phosphate termini and for tandem synthesis.

Main Results:

Demonstrated successful oligonucleotide synthesis using inexpensive LCAA-CPG supports without modifications to standard coupling or dephosphorylation steps.
Achieved the production of oligonucleotides exclusively with terminal 3'-OH ends.
Successfully synthesized linked oligonucleotides via tandem synthesis, yielding mixtures of products upon deprotection.
Showcased the utility for producing pairs of PCR primers or both strands of double-stranded DNA in a single process.

Conclusions:

Developed a versatile and cost-effective approach for oligonucleotide synthesis using novel phosphoramidite reagents and underivatized supports.
The method simplifies the synthesis process, eliminating the need for 3'-dephosphorylation and allowing for tandem synthesis.
This technique offers significant advantages for applications requiring multiple oligonucleotide sequences, such as PCR primer generation and double-stranded DNA synthesis.