Long, Processive Enzymatic Dna Synthesis Using 100% Dye-Labeled Terminal Phosphate-Linked Nucleotides

We demonstrate the efficient synthesis of DNA with complete replacement of the four deoxyribonucleoside triphosphate (dNTP) substrates with nucleotides carrying fluorescent labels. A different, spectrally separable fluorescent dye suitable for single molecule fluorescence detection was conjugated to...

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Detalles Bibliográficos
Autores principales: Korlach, Jonas, Bibillo, Arek, Wegener, Jeffrey, Peluso, Paul, Pham, Thang T., Park, Insil, Clark, Sonya, Otto, Geoff A., Turner, Stephen W.
Formato: Texto
Lenguaje:English
Publicado: Taylor & Francis 2008
Materias:
Original Article
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2582155/
https://www.ncbi.nlm.nih.gov/pubmed/18711669
http://dx.doi.org/10.1080/15257770802260741
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author Korlach, Jonas
Bibillo, Arek
Wegener, Jeffrey
Peluso, Paul
Pham, Thang T.
Park, Insil
Clark, Sonya
Otto, Geoff A.
Turner, Stephen W.
author_facet Korlach, Jonas
Bibillo, Arek
Wegener, Jeffrey
Peluso, Paul
Pham, Thang T.
Park, Insil
Clark, Sonya
Otto, Geoff A.
Turner, Stephen W.
author_sort Korlach, Jonas
collection PubMed
description We demonstrate the efficient synthesis of DNA with complete replacement of the four deoxyribonucleoside triphosphate (dNTP) substrates with nucleotides carrying fluorescent labels. A different, spectrally separable fluorescent dye suitable for single molecule fluorescence detection was conjugated to each of the four dNTPs via linkage to the terminal phosphate. Using these modified nucleotides, DNA synthesis by ø29 DNA polymerase was observed to be processive for products thousands of bases in length, with labeled nucleotide affinities and DNA polymerization rates approaching unmodified dNTP levels. Results presented here show the compatibility of these nucleotides for single-molecule, real-time DNA sequencing applications.
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spelling pubmed-25821552008-11-14 Long, Processive Enzymatic Dna Synthesis Using 100% Dye-Labeled Terminal Phosphate-Linked Nucleotides Korlach, Jonas Bibillo, Arek Wegener, Jeffrey Peluso, Paul Pham, Thang T. Park, Insil Clark, Sonya Otto, Geoff A. Turner, Stephen W. Nucleosides Nucleotides Nucleic Acids Original Article We demonstrate the efficient synthesis of DNA with complete replacement of the four deoxyribonucleoside triphosphate (dNTP) substrates with nucleotides carrying fluorescent labels. A different, spectrally separable fluorescent dye suitable for single molecule fluorescence detection was conjugated to each of the four dNTPs via linkage to the terminal phosphate. Using these modified nucleotides, DNA synthesis by ø29 DNA polymerase was observed to be processive for products thousands of bases in length, with labeled nucleotide affinities and DNA polymerization rates approaching unmodified dNTP levels. Results presented here show the compatibility of these nucleotides for single-molecule, real-time DNA sequencing applications. Taylor & Francis 2008-08-18 2008-09 /pmc/articles/PMC2582155/ /pubmed/18711669 http://dx.doi.org/10.1080/15257770802260741 Text en © Taylor & Francis http://creativecommons.org/licenses/by/2.0/ This is an open access article distributed under the Supplemental Terms and Conditions for iOpenAccess articles published in Taylor & Francis journals (http://www.informaworld.com/mpp/uploads/iopenaccess_tcs.pdf) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Original Article
Korlach, Jonas
Bibillo, Arek
Wegener, Jeffrey
Peluso, Paul
Pham, Thang T.
Park, Insil
Clark, Sonya
Otto, Geoff A.
Turner, Stephen W.
Long, Processive Enzymatic Dna Synthesis Using 100% Dye-Labeled Terminal Phosphate-Linked Nucleotides
title Long, Processive Enzymatic Dna Synthesis Using 100% Dye-Labeled Terminal Phosphate-Linked Nucleotides
title_full Long, Processive Enzymatic Dna Synthesis Using 100% Dye-Labeled Terminal Phosphate-Linked Nucleotides
title_fullStr Long, Processive Enzymatic Dna Synthesis Using 100% Dye-Labeled Terminal Phosphate-Linked Nucleotides
title_full_unstemmed Long, Processive Enzymatic Dna Synthesis Using 100% Dye-Labeled Terminal Phosphate-Linked Nucleotides
title_short Long, Processive Enzymatic Dna Synthesis Using 100% Dye-Labeled Terminal Phosphate-Linked Nucleotides
title_sort long, processive enzymatic dna synthesis using 100% dye-labeled terminal phosphate-linked nucleotides
topic Original Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2582155/
https://www.ncbi.nlm.nih.gov/pubmed/18711669
http://dx.doi.org/10.1080/15257770802260741
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