Abstract
Double-stranded DNA (dsDNA) templates can hybridize to and accelerate cleavage of oligonucleotides containing a P3’→N5’ phosphoramidate (P-N) linkage. This dsDNA-templated cleavage of P-N linkages could be due to conformational strain placed on the linkage upon triplex formation. To determine whether duplex formation also induced conformational strain, we examined the reactivity of the oligonucleotides with a P-N linkage in the presence of single-stranded templates, and compared these reactions to those with dsDNA templates. P-N oligonucleotides that are cleaved upon duplex formation could be used as probes to detect single-stranded nucleic acids.
Highlights
We presented a new type of DNA-templated reaction-based DNA detection; this method involves probe cleavage that is templated by double-stranded DNA [26,27,28,29]
We hypothesized that duplex formation, like triplex formation, could induce conformational strain when oligonucleotides have a certain chemical modification and that such oligonucleotides may be selectively cleaved in the presence of single-stranded templates and, may be used as probes to detect single-stranded nucleic acids (Figure 2)
We evaluated the ability of ON-1–ON-8 to form duplexes with oligonucleotide templates
Summary
Detection of nucleic acids via nucleic acid-templated reactions has attracted substantial interest [1,2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18,19,20,21,22,23,24]. We presented a new type of DNA-templated reaction-based DNA detection; this method involves probe cleavage that is templated by double-stranded DNA (dsDNA) [26,27,28,29] In this method, cleavage reactions are accelerated because of conformational strain induced by triplex formation, not because of the effective molecularity. We examined the effects of chemical modifications that alter the microenvironment around the P-N linkage and change the extent of the conformational strain; these chemical modifications had substantial effects on the observed pseudo first-order rate constants (kobss) of the hydrolysis with the dsDNA templates [29]. We hypothesized that duplex formation, like triplex formation, could induce conformational strain when oligonucleotides have a certain chemical modification and that such oligonucleotides may be selectively cleaved in the presence of single-stranded templates and, may be used as probes to detect single-stranded nucleic acids (Figure 2). Template nucleic acids can be dsDNA or parallel or anti-parallel single-stranded nucleic acids
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