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Fluoride-cleavable, fluorescently labelled reversible terminators: synthesis and use in primer extension.

Fluorescent 2′‐deoxynucleotides containing a protecting group at the 3′‐O‐position are reversible terminators that enable array‐based DNA sequencing‐by‐synthesis (SBS) approaches. Herein, we describe the synthesis and full characterisation of four reversible terminators bearing a 3′‐blocking moiety... Full description

Journal Title: Chemistry (Weinheim an der Bergstrasse Germany), March 1, 2011, Vol.17(10), pp.2903-2915
Main Author: Knapp, Diana C
Other Authors: Serva, Saulius , D'Onofrio, Jennifer , Keller, Angelika , Lubys, Arvydas , Kurg, Ants , Remm, Maido , Engels, Joachim W
Format: Electronic Article Electronic Article
Language: English
Subjects:
ID: E-ISSN: 1521-3765 ; DOI: 10.1002/chem.201001952
Link: http://search.proquest.com/docview/853678641/?pq-origsite=primo
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recordid: proquest853678641
title: Fluoride-cleavable, fluorescently labelled reversible terminators: synthesis and use in primer extension.
format: Article
creator:
  • Knapp, Diana C
  • Serva, Saulius
  • D'Onofrio, Jennifer
  • Keller, Angelika
  • Lubys, Arvydas
  • Kurg, Ants
  • Remm, Maido
  • Engels, Joachim W
subjects:
  • DNA Primers–Metabolism
  • Fluorescent Dyes–Chemical Synthesis
  • Fluorides–Chemistry
  • Leukemia Virus, Murine–Chemistry
  • Molecular Structure–Enzymology
  • Oligonucleotide Probes–Chemical Synthesis
  • RNA-Directed DNA Polymerase–Chemistry
  • RNA-Directed DNA Polymerase–Metabolism
  • DNA Primers
  • Fluorescent Dyes
  • Oligonucleotide Probes
  • RNA-Directed DNA Polymerase
  • Fluorides
ispartof: Chemistry (Weinheim an der Bergstrasse, Germany), March 1, 2011, Vol.17(10), pp.2903-2915
description: Fluorescent 2′‐deoxynucleotides containing a protecting group at the 3′‐O‐position are reversible terminators that enable array‐based DNA sequencing‐by‐synthesis (SBS) approaches. Herein, we describe the synthesis and full characterisation of four reversible terminators bearing a 3′‐blocking moiety and a linker‐dye system that is removable under the same fluoride‐based treatment. Each nucleotide analogue has a different fluorophore attached to the base through a fluoride‐cleavable linker and a 2‐cyanoethyl moiety as the 3′‐blocking group, which can be removed by using a fluoride treatment as well. Furthermore, we identified a DNA polymerase, namely, RevertAid M‐MuLV reverse transcriptase, which can incorporate the four modified reversible terminators. The synthesised nucleotides and the optimised DNA polymerase were used on CodeLink slides spotted with hairpin oligonucleotides to demonstrate their potential in a cyclic reversible terminating approach. Bright...
language: eng
source:
identifier: E-ISSN: 1521-3765 ; DOI: 10.1002/chem.201001952
fulltext: fulltext
issn:
  • 15213765
  • 1521-3765
url: Link


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titleFluoride-cleavable, fluorescently labelled reversible terminators: synthesis and use in primer extension.
creatorKnapp, Diana C ; Serva, Saulius ; D'Onofrio, Jennifer ; Keller, Angelika ; Lubys, Arvydas ; Kurg, Ants ; Remm, Maido ; Engels, Joachim W
contributorKnapp, Diana C (correspondence author) ; Knapp, Diana C (record owner)
ispartofChemistry (Weinheim an der Bergstrasse, Germany), March 1, 2011, Vol.17(10), pp.2903-2915
identifierE-ISSN: 1521-3765 ; DOI: 10.1002/chem.201001952
subjectDNA Primers–Metabolism ; Fluorescent Dyes–Chemical Synthesis ; Fluorides–Chemistry ; Leukemia Virus, Murine–Chemistry ; Molecular Structure–Enzymology ; Oligonucleotide Probes–Chemical Synthesis ; RNA-Directed DNA Polymerase–Chemistry ; RNA-Directed DNA Polymerase–Metabolism ; DNA Primers ; Fluorescent Dyes ; Oligonucleotide Probes ; RNA-Directed DNA Polymerase ; Fluorides
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descriptionFluorescent 2′‐deoxynucleotides containing a protecting group at the 3′‐O‐position are reversible terminators that enable array‐based DNA sequencing‐by‐synthesis (SBS) approaches. Herein, we describe the synthesis and full characterisation of four reversible terminators bearing a 3′‐blocking moiety and a linker‐dye system that is removable under the same fluoride‐based treatment. Each nucleotide analogue has a different fluorophore attached to the base through a fluoride‐cleavable linker and a 2‐cyanoethyl moiety as the 3′‐blocking group, which can be removed by using a fluoride treatment as well. Furthermore, we identified a DNA polymerase, namely, RevertAid M‐MuLV reverse transcriptase, which can incorporate the four modified reversible terminators. The synthesised nucleotides and the optimised DNA polymerase were used on CodeLink slides spotted with hairpin oligonucleotides to demonstrate their potential in a cyclic reversible terminating approach. Bright...
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