This modification is a post synthesis maleimide conjugation to a reduced thiol amino group thus an additional modification with thiol group is required. A C3 or C6 thiol group can be placed at the 5' or for internal positions Thiol C6 dT modified base is used.
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Azide PEG3 Maleimide can be used to introduce an active azide group to a thiol-modified oligonucleotide. The Azide C2 NHS ester is then manually attached to the oligo through the amino group in a separate reaction post-synthesis.
The presence of the azide allows the user to use "Click Chemistry" (a [3+2] cycloaddition reaction between alkynes and azides, using copper (I) iodide as a catalyst) to conjugate the azide-modified oligo to a terminal alkyne-modified oligo with extremely high regioselectivity and efficiency (1,2). Preparation of the alkyne-modified oligo can be achieved using the 5’-Hexynyl modifier (see its respective tech sheet for details). Click chemistry can be used to form short, cyclic oligos that can be used as research tools in various biophysical and biological studies (3). In particular, they have considerable potential for in vivo work, as cyclic oligos are known to be very stable in serum for up to several days.
1. Huisgen, R. Angew. Chem. Int. Ed. (1963), 2: 565-568.
2. Rostovtsev, V.V., Green, L.G., Fokin, V.V., Sharpless, K.B. A Stepwise Huisgen Cycloaddition Process: Copper(I)-Catalyzed Regioselective Ligation of Azides and Terminal Alkynes. Angew. Chem. Int. Ed. (2002), 41: 2596-2599.
3. Kumar, R., El-Sagheer, A., Tumpane, J., Lincoln, P., Wilhelmsson, L.M., Brown, T. Template-Directed Oligonucleotide Strand Ligation, Covalent Intramolecular DNA Circularization and Catenation Using Click Chemistry. J. Am. Chem. Soc. (2007), 129: 6859-6864.