SUMMARY. Experimental comparison of fluorescent quenching efficacy for two oligonucleotide probes in different conformations such as molecular beacon probe and TaqMan linear probe was performed. Interaction of quencher with fluorophore is different because these probes have different conformations. For linear probe quenching arises by inductive Forster resonance ener-gy transfer (FRET) mechanism. For hairpin probe quenching occurs through contact quenching by more close location of fluorophore and quencher but Forster resonance energy transfer may occur too. Absorbance spectrum of linear probe practically coincides with spectrum oligonucleotide which represent probe without quencher that indicates onto dynamical (Forster) resonance energy transfer mechanism. Contrary, absorbance spectra of hairpin probe and oligonucleotide which represent probe without quencher are significantly different that points to contact energy transfer mechanism between fluorophore and quencher. Fluorescence spectra of probes and their complexes with complementary to linear probe oligonucleotide (and to loop of hairpin probe) and with amplicon (with length of 200 bp and which contains DNA target for probes) allow to compare these two probes by comparing radius of energy migration, fluorescent quenching efficacy. Radius of energy migration (R) calculated from expe-rimental data for hairpin probe was 32,4 Å and for linear probe was 47,3 Å. Hairpin probe in comparison with linear one is characterized by higher efficacy of fluorescent quenching that results in lower fluorescence background and therefore higher level of signal/noise during RT-PCR.
Ключові слова: гасіння флуоресценції, флуоресцентний зонд, характеризація зонда, ПЛР у реальному часі, ФРПЕ, зв’язування олігонуклеотида, олігонуклеотидний дуплекс, спектри флуоресценції
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