To broaden the scope of applications in DNA nano- and biotechnology, material science, diagnostics and molecular recognition the functionalization of DNA is of utmost importance. In the last decade many new methods have been developed to achieve this goal. Apart from the direct chemical synthesis of modified DNA by automated phosphoramidite chemistry incorporation of labelled triphosphates and the post-synthetic labelling approach evolved as valuable methods. New bioorthogonal reactions as Diels-Alder, click and Staudinger ligations pushed forward the post-synthetic approach as new insights into DNA polymerase substrate specificity allowed generation and amplification of labelled DNA strands. These novel developments are summarized herein.

A novel nucleotide building block for enzymatic synthesis of azide modified DNA and subsequent conjugation via Staudinger ligation was developed.Many studies of complex biological systems are only feasible through conjugation of biopolymers with other functional molecules like dyes or affinity tags. Linkage of a complex biomolecule with another molecular entity is often a challenging task. Albeit several bioconjugation methods are available, truly chemoselective methods are rarely represented. 1,2 Recently, two reactions based on the specific reactivity of the azide moiety were introduced. [3][4][5] One method is based on the [3 + 2] cyclo addition reaction of an azide and a terminal alkyne 6 promoted by copper catalysis. 7,8 This reaction has been employed extensively and recently been extended for conjugation of DNA. 9,10 On the other side, the so-called Staudinger ligation developed by Bertozzi [11][12][13] readily occurs between an azide and a phosphine to form an azaylide that can be trapped by an acyl group to form a stable amide bond. This reaction has been employed in numerous applications e.g. for the conjugation of carbohydrates, protein 14,15 and phage particles 16 as well as peptide ligation and immobilization. [17][18][19][20][21] Recently, methods were devised to employ the Staudinger ligation for conjugation of DNA. It was shown that conjugation of an azide functionality via a suitable linker system enables conjugation of dyes to the 59-end of single stranded DNA. 22 Rajski et al. employed the Staudinger ligation to conjugate DNA with phenanthroline moieties for subsequent Cu(I) induced strand scission. 23 Here we report on the development of a novel azide modified nucleoside triphosphate building block, that is readily incorporated into DNA enzymatically by a DNA polymerase. The resulting double stranded azide modified DNA can be smoothly conjugated with modified phosphines via the Staudinger ligation.Our strategy to modify DNA site-specifically with azide functions by use of DNA polymerases for subsequent Staudinger ligation is depicted in Scheme 1B. The first step consists of a DNA polymerase reaction in which one of the natural nucleoside triphosphates (dNTPs) is substituted by a modified analogue that contains an azide functionality. Obviously the success of this step is contingent on the ability of a DNA polymerase to accept the modified nucleotide. The azide-modified double stranded DNA in turn should serve as substrate for Staudinger ligation with a suitably functionalized phosphine.Thus, first we developed the synthesis of the nucleoside triphosphate 1, a 29-deoxyadenosine analogue (Scheme 1A). Since it has been shown that 7-modified 7-deaza-29-deoxyadenosine derivatives are accepted by DNA polymerases 24-28 we focused our synthesis on these kinds of analogues. Following a published procedure 24 we synthesised a 7-deaza-29-deoxyadenosine 2, which was readily converted into the corresponding triphosphate 3. After saponification 24 the azide moiety was introduced via EDC promoted amide bond...

Totally wired: The folding of G‐quadruplexes leads to a stacking of the G‐bases, which provide a relatively large conductance. Stretching these complexes results in changes to the resistance that are only slightly over a broad range of molecular lengths. G‐quadruplexes can thus be seen as molecular wires with adjustable length.

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Two 5 modified 2'-deoxyuridin triphosphates and a 7 modified 2'-deoxy-7-deazaadenosine were synthesized carrying a terminal azide linked to the base.