ArticleProceedings of the National Academy of Sciences of the United States of America2008
Four-color DNA sequencing with 3'-O-modified nucleotide reversible terminators and chemically cleavable fluorescent dideoxynucleotides.
Article in Proceedings of the National Academy of Sciences of the United States of America, 2008. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 62 papers.
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Who cites it
62 citing papers in PubMed.
- Semi-rational engineering of terminal deoxynucleotidyl transferase for high-efficiency enzymaticSynthetic and systems biotechnology · 2026Article
- From liquid-phase synthesis to chemical ligation: preparation of oligonucleotides and their backbone analogs in solution.Nucleic acids research · 2025Review
- Advancing synthesis-free and enzyme-free rewritable DNA memory through frameshift encoding and nanopore duplex interruption decoding.PNAS nexus · 2025Article
- Oligonucleotide subsets selection by single nucleotide resolution barcode identification.Nature communications · 2025Article
- Semi-rational evolution of a recombinant DNA polymerase for modified nucleotide incorporation efficiency.PloS one · 2025Article
- The Evolution of Next-Generation Sequencing Technologies.Methods in molecular biology (Clifton, N.J.) · 2025Review
- From Tradition to Innovation: Diverse Molecular Techniques in the Fight Against Infectious Diseases.Diagnostics (Basel, Switzerland) · 2024Review
- Controlled enzymatic synthesis of oligonucleotides.Communications chemistry · 2024Review
- A New Fluorescent Probe for Hydrogen Sulfide Detection in Solution and Living Cells.Molecules (Basel, Switzerland) · 2023Article
- A new method to synthesize multiple gRNA libraries and functional mapping of mammalian H3K4me3 regions.Nucleic acids research · 2023Article
- The Principles and Applications of High-Throughput Sequencing Technologies.Development & reproduction · 2023Article
- Review
- Towards the controlled enzymatic synthesis of LNA containing oligonucleotides.Frontiers in chemistry · 2023Article
- Application of next-generation sequencing to identify different pathogens.Frontiers in microbiology · 2023Review
- DNA synthesis technologies to close the gene writing gap.Nature reviews. Chemistry · 2023Review
- Evaluation of the correctable decoding sequencing as a new powerful strategy for DNA sequencing.Life science alliance · 2022Article
- Evaluation of 3'-phosphate as a transient protecting group for controlled enzymatic synthesis of DNA and XNA oligonucleotides.Communications chemistry · 2022Article
- Design considerations for advancing data storage with synthetic DNA for long-term archiving.Materials today. Bio · 2022Review
- Application and Challenge of 3rd Generation Sequencing for Clinical Bacterial Studies.International journal of molecular sciences · 2022Review
- Purification of multiplex oligonucleotide libraries by synthesis and selection.Nature biotechnology · 2022Article
2 more citing papers are in PubMed but not listed here.
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Authors and funding
16 authors.
Funding
Abstract
DNA sequencing by synthesis (SBS) on a solid surface during polymerase reaction can decipher many sequences in parallel. We report here a DNA sequencing method that is a hybrid between the Sanger dideoxynucleotide terminating reaction and SBS. In this approach, four nucleotides, modified as reversible terminators by capping the 3'-OH with a small reversible moiety so that they are still recognized by DNA polymerase as substrates, are combined with four cleavable fluorescent dideoxynucleotides to perform SBS. The ratio of the two sets of nucleotides is adjusted as the extension cycles proceed. Sequences are determined by the unique fluorescence emission of each fluorophore on the DNA products terminated by ddNTPs. On removing the 3'-OH capping group from the DNA products generated by incorporating the 3'-O-modified dNTPs and the fluorophore from the DNA products terminated with the ddNTPs, the polymerase reaction reinitiates to continue the sequence determination. By using an azidomethyl group as a chemically reversible capping moiety in the 3'-O-modified dNTPs, and an azido-based cleavable linker to attach the fluorophores to the ddNTPs, we synthesized four 3'-O-azidomethyl-dNTPs and four ddNTP-azidolinker-fluorophores for the hybrid SBS. After sequence determination by fluorescence imaging, the 3'-O-azidomethyl group and the fluorophore attached to the DNA extension product via the azidolinker are efficiently removed by using Tris(2-carboxyethyl)phosphine in aqueous solution that is compatible with DNA. Various DNA templates, including those with homopolymer regions, were accurately sequenced with a read length of >30 bases by using this hybrid SBS method on a chip and a four-color fluorescence scanner.
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