Abstract
Site-specific and chemoselective labeling of DNA is still a difficult task. The Staudinger ligation is a bioorthogonal reaction between azides and phosphines that requires no catalyst to proceed, allowing for mild reaction conditions. The reaction may be extended for site-specific labeling of DNA using azido-modified triphosphates, which can be incorporated site-specifically into DNA strands by DNA polymerases in a template-dependent manner. The azido-modified DNA, in turn, can be labeled by suitable phosphines. This protocol describes (1) the synthesis of an azido-TTP analogue; (2) the enzymatic synthesis of azido-modified DNA; (3) the synthesis of suitable phosphine labels; and (4) the labeling of azido-DNA with biotin–phosphine by Staudinger ligation with approximately 70% conversion.
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References
Niemeyer, C. M. ed. (2004) Bioconjugation Protocols (Methods in Molecular Biology Series Vol. 283). Humana Press, Totowa, NJ.
Weisbrod, S. H. and Marx, A. (2008) Novel strategies for the site-specific covalent labeling of nucleic acids. Chem. Commun. 44, 5675–5685.
Saxon, E. and Bertozzi, C. R. (2000) Cell surface engineering by a modified Staudinger reaction. Science 287, 2007–2010.
Prescher, J. A., Dube D. H. and Bertozzi C. R. (2004) Chemical remodelling of cell surfaces in living animals. Nature 430, 873–877.
Weisbrod, S. H. and Marx, A. (2007) A nucleoside triphosphate for site-specific labeling of DNA by the Staudinger ligation. Chem. Commun. 18, 1828–1830.
Baccaro, A. Weisbrod, S. H. and Marx, A. (2007) DNA conjugation by the Staudinger ligation: new thymidine analogues. Synthesis 13, 1949–1954.
Robins, M. J. and Barr, P. J. (1983) Nucleic acid related compounds. 39. Efficient conversion of 5-iodo to 5-alkynyl and derived 5-substituted uracil bases and nucleosides. J. Org. Chem. 48, 1854–1862.
Sambrook J., Russell D. W. (2001) Molecular Cloning: A Laboratory Manual. Laboratory Press, Cold Spring Harbor, NY.
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Weisbrod, S.H., Baccaro, A., Marx, A. (2011). Site-Specific DNA Labeling by Staudinger Ligation. In: Mark, S. (eds) Bioconjugation Protocols. Methods in Molecular Biology, vol 751. Humana Press. https://doi.org/10.1007/978-1-61779-151-2_12
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DOI: https://doi.org/10.1007/978-1-61779-151-2_12
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