title: Rapid genotyping using pyrene-perylene locked nucleic acid complexes.
abstract: We have developed an assay for single strand DNA and RNA detection which is based on novel pyrene-perylene FRET pairs attached to short LNA/DNA probes. The assay is based on ratiometric emission upon binding of target DNA/RNA by three combinations of fluorescent LNA/DNA reporter strands. Specific geometry of the pyrene fluorophore attached to the 2'-amino group of 2'-amino-LNA in position 4 allows for the first time to efficiently utilize dipole-dipole orientation parameter for sensing of single-nucleotide polymorphisms (SNPs) in nucleic acid targets by FRET. Using novel probes, SNP detection is achieved with advantages of large Stokes shift (115 nm), high fluorescence quantum yields and low limit of target detection values (< 5 nM). Rapid and accurate genotyping of highly polymorphic HIV Pol cDNA and RNA fragments performed herein proves the possibility for broad application of the novel pyrene-perylene FRET pairs, e.g., in imaging and clinical diagnostics.
authors.affiliation: Nucleic Acid Center; Department of Physics, Chemistry and Pharmacy; University of Southern Denmark; Odense, Denmark ; National Institute of Diabetes and Digestive and Kidney Diseases; National Institutes of Health; Molecular Recognition Section; Bethesda, MD USA.
authors.forename: T Santhosh
authors.initials: TS
authors.lastname: Kumar
authors.forename: Anna
authors.initials: A
authors.lastname: Myznikova
authors.forename: Evgeniya
authors.initials: E
authors.lastname: Samokhina
authors.forename: Irina Kira
authors.initials: IK
authors.lastname: Astakhova
mesh_terms: D001483:Base Sequence; D015342:DNA Probes / Q000737:chemistry / Q000235:genetics; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes / Q000737:chemistry*; D005838:Genotype; D060005:Genotyping Techniques; D006801:Humans; D057230:Limit of Detection; D009841:Oligonucleotides / Q000737:chemistry / Q000235:genetics*; D010569:Perylene / Q000737:chemistry*; D020641:Polymorphism, Single Nucleotide; D011721:Pyrenes / Q000737:chemistry*
chemical_list: D015342:DNA Probes; D005456:Fluorescent Dyes; D009841:Oligonucleotides; D011721:Pyrenes; C477371:locked nucleic acid; D010569:Perylene
keywords: FRET; LNA; SNP; fluorescence; oligonucleotide; perylene; pyrene
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