Fast and easy colorimetric tests for single mismatch recognition by PNA-DNA duplexes with the diethylthiadicarbocyanine dye and succinyl-beta-cyclodextrin.
The 3,3'-diethylthiacarbocyanine (DiSC(2)(5)) dye is able to aggregate on full matched PNA-DNA duplexes by changing its absorption properties, which are manifested by an instantaneous colour shift from blue to purple. However the spontaneous aggregation of the dye also on mismatched duplexes and even on free PNA strands makes the test quite aspecific. Here it is demonstrated that the addition of succinyl-beta-cyclodextrin (Succ-beta-CyD) to the solutions containing PNA-DNA duplexes and the dye strongly enhances the specificity of the colour shift, allowing for a fast, very specific and extremely sensitive visual recognition of mismatches in DNA strands by using PNA probes in combination with the DiSC(2)(5) dye. The phenomenon has been studied by CD and NMR spectroscopies. The method has been optimized and preliminarily applied for the recognition of an apoE gene mutation in human DNA samples.
Tedeschi
Tullia
T
Department of Organic and Industrial Chemistry, University of Parma, Viale G.P. Usberti 17/A, I-43100, Parma, Italy.
Sforza
Stefano
S
Ye
Sheng
S
Corradini
Roberto
R
Dossena
Arnaldo
A
Komiyama
Makoto
M
Marchelli
Rosangela
R
D001057:Apolipoproteins E / Q000235:genetics; D020137:Base Pair Mismatch; D001483:Base Sequence; D003124:Colorimetry / Q000379:methods*; D004396:Coloring Agents; D004247:DNA / Q000737:chemistry* / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D004225:Dithiazanine; D005456:Fluorescent Dyes; D006801:Humans; D066298:In Vitro Techniques; D046911:Macromolecular Substances / Q000737:chemistry; D008969:Molecular Sequence Data; D020135:Peptide Nucleic Acids / Q000737:chemistry*; D017354:Point Mutation; D020641:Polymorphism, Single Nucleotide; D012997:Solvents; D013053:Spectrophotometry; D047392:beta-Cyclodextrins / Q000737:chemistry
D001057:Apolipoproteins E; D004396:Coloring Agents; D017931:DNA Primers; D005456:Fluorescent Dyes; D046911:Macromolecular Substances; D020135:Peptide Nucleic Acids; D012997:Solvents; D047392:beta-Cyclodextrins; D004225:Dithiazanine; D004247:DNA