DNA with 3′‐5′‐Disulfide Links—Rapid Chemical Ligation through Isosteric Replacement |
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| Authors: | Dr Volker Patzke Prof?Dr John S McCaskill Prof?Dr Günter von?Kiedrowski |
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| Affiliation: | 1. Lehrstuhl für Bioorganische Chemie, Ruhr‐Universit?t Bochum, 44780 Bochum (Germany);2. BioMIP: Microsystems Chemistry and BioIT, Ruhr‐Universit?t Bochum, 44780 Bochum (Germany) |
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| Abstract: | Efforts to chemically ligate oligonucleotides, without resorting to biochemical enzymes, have led to a multitude of synthetic analogues, and have extended oligomer ligation to reactions of novel oligonucleotides, peptides, and hybrids such as PNA. 1 Key requirements for potential diagnostic tools not based on PCR include a fast templated chemical DNA ligation method that exhibits high pairing selectivity, and a sensitive detection method. Here we report on a solid‐phase synthesis of oligonucleotides containing 5′‐ or 3′‐mercapto‐dideoxynucleotides and their chemical ligations, yielding 3′‐5′‐disulfide bonds as a replacement for 3′‐5′‐phosphodiester units. Employing a system designed for fluorescence monitoring, we demonstrate one of the fastest ligation reactions with half‐lives on the order of seconds. The nontemplated ligation reaction is efficiently suppressed by the choice of DNA modification and the 3′‐5′ orientation of the activation site. The influence of temperature on the templated reaction is shown. |
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| Keywords: | chemical ligation FRET kinetics oligonucleotides sulfur |
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