Chemical reactions within DNA duplexes Cyanogen bromide as an effective oligodeoxyribonucleotide coupling agent

Abstract

Cyanogen bromide was found to condense oligodeoxyribonucleotides on a complementary template in aqueous solution. Optimum conditions for this vigorous and effective reaction were developed. CNBr proved to be useful for incorporation of phosphoramidate or pyrophosphate internucleotide bonds in DNA duplexes.

Biocompatible artificial DNA linker that is read through by DNA polymerases and is functional in Escherichia coli

“…In addition, some modified bases are not tolerated by ligase enzymes. Enzymatic methods of gene synthesis are extremely important in biology, but a purely chemical method for the assembly of large DNA molecules would be an interesting and valuable addition to current tools, with the advantages of scalability, flexibility, and orthogonality.It has proved challenging to achieve clean and efficient chemical ligation of canonical DNA, although significant progress has been made using cyanogen bromide as a coupling agent (11,12). An interesting alternative approach is to design a chemical linkage that mimics the natural phosphodiester bond, and that can be formed in high yield in aqueous media from functional groups that are orthogonal to those naturally present in DNA.…”

“…It has proved challenging to achieve clean and efficient chemical ligation of canonical DNA, although significant progress has been made using cyanogen bromide as a coupling agent (11,12). An interesting alternative approach is to design a chemical linkage that mimics the natural phosphodiester bond, and that can be formed in high yield in aqueous media from functional groups that are orthogonal to those naturally present in DNA.…”

Preparation of modified long-mer RNAs and analysis of FMN binding to theypaAaptamer fromB. subtilis

“…In addition to enzymatic protocols for ligation of chemically synthesized fragments, chemical protocols were developed, [40][41][42][43] whereby the cupper-catalyzed azide-alkyne coupling (Click ligation) 26,29,34,44 stands out as most promising. A drawback of Click ligation is the need for terminal functionalization of fragments to be ligated (5′-azide and 3′-alkyne or vice versa), and the generation of a triazole linkage in the nucleic acid backbone.…”

“…Nevertheless, Click ligation has been used for fragment joining in a number of applications. 26,29,34,[40][41][42][43][44] Among those, also functional RNAs, such as hairpin and hammerhead ribozymes, were prepared. 29 However, to the best of our knowledge, we here show the first example of a Click ligated natural aptamer as long as 129 nucleotides that retains full functionality in binding of its cognate ligand, in spite of the two triazole backbone linkages.…”

A triazole linkage that mimics the DNA phosphodiester group in living systems

“…Circumventing the problem by assembling DNA from purified short oligonucleotides will only serve to increase the number of ligation steps that are necessary. This is a concern, because the phosphodiester bonds between oligonucleotide strands have proved to be difficult to create chemically in high yield, although some progress has been made using cyanogen bromide as a coupling agent (Luebke & Dervan, 1991;Sokolova et al 1987). An alternative strategy is to design an artificial chemical linkage that mimics the natural phosphodiester bond and which can be formed in high yield in aqueous media from functional groups that are orthogonal to those present in DNA.…”