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Comparative Study
. 2017 Nov;27(6):2001-2016.
doi: 10.1007/s10895-017-2139-6. Epub 2017 Jul 28.

Comparative Study of Novel Fluorescent Cyanine Nucleotides: Hybridization Analysis of Labeled PCR Products Using a Biochip

V E Shershov  1 , S A Lapa  1 , V E Kuznetsova  2 , M A Spitsyn  1 , T O Guseinov  1 , S A Polyakov  1 , A A Stomahin  1 , A S Zasedatelev  1 , A V Chudinov  1
Affiliations
Comparative Study

Comparative Study of Novel Fluorescent Cyanine Nucleotides: Hybridization Analysis of Labeled PCR Products Using a Biochip

V E Shershov et al. J Fluoresc. 2017 Nov.

Abstract

This study investigated the synthesis and substrate properties of Cy5-labeled dUTP derivatives with different substituents, linkers between the dye unit and pyrimidine heterocycle and fluorophore charges. Fluorescently labeled nucleoside triphosphates were studied as substrates using multiplex PCR with Taq and Vent (exo-) DNA polymerases, the typical representatives of the A and B polymerase families. The efficiency of nucleotide incorporation during PCR was assessed with a multi-parameter hybridization analysis using a diagnostic DNA microarray. The hybridization analysis indirectly estimates the incorporation efficiency of dye-labeled nucleotides in multiplex PCR. Our results demonstrated higher efficiencies of substrates with electrically neutral dyes than electropositive and electronegative Cy5 residues.

Keywords: Biochip; Cyanine dyes; Fluorescently labeled nucleotides; Fluorophore charge; PCR.

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