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PMID: 18263613 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

A new class of cleavable fluorescent nucleotides: synthesis and optimization as reversible terminators for DNA sequencing by synthesis.

Nucleic acids research ·Vol. 36 ·No. 4 ·2008-03-00 ·Pages e25

Turcatti G, Romieu A, Fedurco M, Tairi AP

Abstract

Fluorescent 2'-deoxynucleotides containing a protecting group at the 3'-O-position are reversible terminators enabling array-based DNA sequencing by synthesis (SBS) approaches. Herein, we describe the synthesis of a new family of 3'-OH unprotected cleavable fluorescent 2'-deoxynucleotides and their evaluation as reversible terminators for high-throughput DNA SBS strategies. In this first version, all four modified nucleotides bearing a cleavable disulfide Alexa Fluor(R) 594 dye were assayed for their ability to act as a reversible stop for the incorporation of the next labeled base. Their use in SBS leaded to a signal-no signal output after successive addition of each labeled nucleotide during the sequencing process (binary read-out). Solid-phase immobilized synthetic DNA target sequences were used to optimize the method that has been applied to DNA polymerized colonies or clusters obtained by in situ solid-phase amplification of fragments of genomic DNA templates.

MeSH Terms
Deoxyribonucleotides/chemical synthesis,chemistry,classification Fluorescent Dyes/chemistry Microscopy, Fluorescence Organic Chemicals/chemistry Sequence Analysis, DNA
Chemicals
Alexa594 Deoxyribonucleotides Fluorescent Dyes Organic Chemicals
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Turcatti Gerardo
Manteia Predictive Medicine S.A., Zone Industrielle, Coinsins, CH-1267, Switzerland. [email protected]
Romieu Anthony
Fedurco Milan
Tairi Ana-Paula
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Article Info
Journal
Nucleic acids research
Abbr.
Nucleic Acids Res
ISSN
1362-4962
DOI
10.1093/nar/gkn021
Email
[email protected]
Published
2008-03-00
Epub
2008-00-07
Pages
e25
Language
English
Region
England
NLM ID
0411011
PMCID
PMC2275100
Subset
IM
External Links
PubMed source DOI: 10.1093/nar/gkn021 PMC Full Text View journal details
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