Synthesis, Dynamic Combinatorial Chemistry, and PCR Amplification of 3′–5′ and 3′–6′ Disulfide‐linked Oligonucleotides

Abstract

Disulfide dithymidines linked 3'-5' or 3'-6' were synthesized and incorporated into oligonucleotides through a combined phosphotriester and phosphoramidite solid-phase oligonucleotide synthesis approach. The disulfide links are cleaved and formed reversibly in the presence of thiols and oligonucleotides. This link was shown to be sequence-adaptive in response to given templates in the presence of mercaptoethanol. The artificial 3'-5' and 3'-6' disulfide link was tolerated by polymerases in the polymerase chain reaction (PCR). By using sequencing analysis, we show that single mutations frequently occurred randomly in the amplification products of the PCR.

Squaramides and Ureas: A Flexible Approach to Polymerase‐Compatible Nucleic Acid Assembly

“…However, a significant limitation is our dependence on ligase enzymes to join oligonucleotide strands together; these enzymes require highly specific reaction conditions and substrates. Consequently purely chemical nucleic acid ligation methods have been explored; amide, CuAAC, phosphoramidate (PA), phosphorothioate (PS), and thiol–thiol coupling reactions generate artificial backbones that are recognised and read‐through by DNA polymerases, thereby enabling information retrieval. Yet all face potential drawbacks, including copper dependence (CuAAC), precursor handling (disulfide, PA, PS), slow ligation rates (PS), and poor read‐through fidelity (disulfide).…”

Self-Assembly Can Direct Dynamic Covalent Bond Formation toward Diversity or Specificity

“… 6 − 8 Noncovalent interactions 9 can then be utilized to channel the building blocks into specific DCL members that optimally engage in molecular recognition. This effect has been exploited for the dynamic combinatorial discovery of synthetic receptors 10 − 17 and ligands for biomolecules 18 , 19 by exposing the system to a corresponding template.…”

Dynamic Chemistry of Disulfide Terminated Oligonucleotides in Duplexes and Double‐Crossover Tiles

“…This study required free thiols for the disulfide formation in all cases and did not take advantage of the reversibility of such connection. We recently showed that a disulfide bond could be integrated in the backbone of a DNA strand and used in a dynamic combinatorial fashion for the templated disproportionation of oligonucleotides . In a related study, Patzke et al.…”

“…We recently showedt hat ad isulfide bond could be integrated in the backbone of aD NA strand and used in ad ynamic combinatorial fashion for the templated disproportionation of oligonucleotides. [39] In ar elateds tudy, Patzke et al demonstrated the ligation of DNA strandsi n which the phosphate linkagew as replaced with ad isulfide. [40] Here we presentamethodt or eversibly connecta djacent DNA strands in af olded structure based on the principleso f DNA-templated synthesis and the dynamic combinatorial chemistry of disulfides ( Figure 1).…”