Issue 35, 2016

Photo-cleavable nucleotides for primer free enzyme mediated DNA synthesis

Abstract

The synthesis, characterization and potential application of eight 3′-O modified 2′-deoxyribonucleoside triphosphates (dNTPs) are discussed. These nucleotide analogues are modified by capping the 3′-OH with a photolabile protecting group which can temporarily cease DNA strand growth and can smoothly reinitiate the growth by the photodecomposition of the protecting group and setting the 3′-OH of dNTPs free to propagate. The synthesis of 3′-O-(2-nitrobenzyl)-2′-deoxyribonucleoside triphosphates (NB-dNTPs) and 3′-O-(4,5-dimethoxy-2-nitrobenzyl)-2′-deoxyribonucleoside triphosphates (DMNB-dNTPs) is discussed in detail with structural confirmation using NMR. The UV-cleaving studies are monitored and quantified using LCMS and 1H NMR spectral traces. The synthesised nucleotides are employed for terminating and reinitiating template-less DNA synthesis, using primer independent Terminal Deoxynucleotidyl Transferase (TdT) enzyme. The use of this photolabile nucleotide in one step stop–start DNA synthesis is a novel strategy towards the precise assembly of dNTPs with the potential to reinforce present technologies.

Graphical abstract: Photo-cleavable nucleotides for primer free enzyme mediated DNA synthesis

Supplementary files

Article information

Article type
Paper
Submitted
24 Jun 2016
Accepted
09 Aug 2016
First published
09 Aug 2016

Org. Biomol. Chem., 2016,14, 8278-8288

Photo-cleavable nucleotides for primer free enzyme mediated DNA synthesis

A. S. Mathews, H. Yang and C. Montemagno, Org. Biomol. Chem., 2016, 14, 8278 DOI: 10.1039/C6OB01371F

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