Fluorescent Color Readout of DNA Hybridization with Thiazole Orange as an Artificial DNA Base

Abstract

A fluorescent chameleon: A single thiazole orange (TO) dye, when used as an artificial DNA base shows the typical green emission, whereas the interstrand TO dimer exhibits an orange excimer-type emission inside duplex DNA (see picture).

Thiazole Orange Dimers in DNA: Fluorescent Base Substitutions with Hybridization Readout

“…Alternatively,T Ow as at-tached to DNA-binding peptides [23] for cell staining or incorporated as ab ase surrogate into peptiden ucleic acid (PNA) [24] as aso-called forced intercalation probe to detect single-base variations. [24b, 25] There are also several reports on the synthesis of TO as aD NA base surrogate by using the nucleoside approach [26] or the approach to substitutet he 2'-deoxyribofuranoside by serinol [27] or (S)-aminopropanediol [28] as an acyclic linker.F or the last case, interstrandT Od imers in DNA show ar emarkable excimer-likes hift of the fluorescencec olor from TO-typical green (l = 530 nm) to orange (l = 580 nm), which can be used as af luorescencer eadout for DNA [29] and RNA hybridization, [30] for imaging of RNA delivery into cells, [30] and for MBs. [29] Herein, we present af ull opticals pectroscopic characterization of the dimerizationofTObase surrogates in different orientations (inter-and intrastrand)a nd different DNA environments.A sm entioned above,b ecause the TO dimer is applicable in any sequential context, it provides ag reat advantage over other fluorophores, especially the PDI dye.…”

“…[29] Herein, we present af ull opticals pectroscopic characterization of the dimerizationofTObase surrogates in different orientations (inter-and intrastrand)a nd different DNA environments.A sm entioned above,b ecause the TO dimer is applicable in any sequential context, it provides ag reat advantage over other fluorophores, especially the PDI dye. [15b, 31] In addition to our preliminary experiments, [29] we show how the TO dimer photophysically interacts within the DNA framework. To representatively demonstrate the use of the fluorescent readout of the TO dimer,M Bs were synthesized to detect singlebase mutations in the target oligonucleotide by fluorescence color changes.…”

Synthetic incorporation of Nile Blue into DNA using 2′-deoxyriboside substitutes: Representative comparison of (R)- and (S)-aminopropanediol as an acyclic linker

“…In comparison to our earlier synthetic protocols for incorporation of ethidium [29–30], indole [31] and phenothiazine[37] the NH group of the carbamate is less nucleophilic and need not be protected during phosphoramidite synthesis. This facilitates the preparation of DNA building blocks as we have shown with indole [32] and thiazole orange [33–34]. DMT-protected ( R )-3-amino-1,2-propanediol 1 as a precursor was synthesized according to literature [29,43].…”

“…( S )-3-Amino-1,2-propanediol was used as an acyclic linker and substitute for the 2′-deoxyriboside between the phosphodiester bridges. Similar propanediol derivatives have been used extensively as alternative and simplified phosphodiester linkers in the 1990s [19–23], and have been further explored for glycol nucleic acid (GNA) [24–26] twisted intercalating nucleic acids (TINA) [27–28], and by our group for fluorescent DNA base substitutions by ethidium [29–30], indole [31–32], thiazole orange [33–34], perylene bisimide [35–36] and phenothiazine [37]. This 2′-deoxyriboside substitution provides high chemical stability and conformational flexibility for the chromophore to intercalate.…”

Fluorescent Nucleic Acid Analogues in Research and Clinical Diagnostics

“…2). Typically, dye intercalation results in increased thermal stability of the double helix and altered fluorescence signal of the dye (Glazer and Rye 1992;Berndl and Wagenknecht 2009). Intercalators can be either attached covalently to the nucleic acid sequence or interact with the duplex non-covalently (Rye and Glazer 1995;Karlsen et al 2012).…”