1269664-56-3Relevant academic research and scientific papers
Single Labeled DNA FIT Probes for Avoiding False-Positive Signaling in the Detection of DNA/RNA in qPCR or Cell Media
Hoevelmann, Felix,Bethge, Lucas,Seitz, Oliver
, p. 2072 - 2081 (2012)
Oligonucleotide hybridization probes that fluoresce upon binding to complementary nucleic acid targets allow the real-time detection of DNA or RNA in homogeneous solution. The most commonly used probes rely on the distance-dependent interaction between a fluorophore and another label. Such duallabeled oligonucleotides signal the change of the global conformation that accompanies duplex formation. However, undesired nonspecific binding events and/or probe degradation also lead to changes in the label-label distance and, thus, to ambiguities in fluorescence signaling. Herein, we introduce singly labeled DNA probes, DNA FIT probes , that are designed to avoid false-positive signals. A thiazole orange (TO) intercalator dye serves as an artificial base in the DNA probe. The probes show little background because the attachment mode hinders 1) interactions of the TO base in cis with the disordered nucleobases of the single strand, and 2) intercalation of the TO nucleotide with double strands in trans. However, formation of the probe-target duplex enforces stacking and increases the fluorescence of the TO base. We explored open-chain and carbocyclic nucleotides. We show that the incorporation of the TO nucleotides has no effect on the thermal stability of the probe-target complexes. DNA and RNA targets provided up to 12-fold enhancements of the TO emission upon hybridization of DNA FIT probes. Experiments in cell media demonstrated that false-positive signaling was prevented when DNA FIT probes were used. Of note, DNA FIT probes tolerate a wide range of hybridization temperature; this enabled their application in quantitative polymerase chain reactions.
Probes comprising fluorescent artificial nucleobases and use thereof for detection of single base alteration
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Page/Page column 13; 14, (2015/12/25)
The present invention relates to forced intercalation probes (FIT-probes) comprising at least one nucleoside analog which comprises at least a fluorescent artificial nucleobase directly bound to a carbon of a modified sugar moiety wherein said modified su
