Integrated DNA Technologies Zen double-quenched probe

Integrated DNA Technologies has introduced a double-quenched probe that increases the accuracy and reliability of 5′ nuclease qPCR experiments. While traditional probes have 20-30 bases between the dye and quencher, this probe design positions an internal Zen quencher only 9 bases from the 5′ fluorophore.

This shortened distance, particularly when combined with the standard 3′ quencher, is claimed to decrease background fluorescence and increase sensitivity. The chemical structure of the Zen quencher stabilises duplex formation, which allows for its use in previously validated sequences. The improved functionality increases qPCR accuracy and sensitivity when compared to traditional probes.

Due to the incorporation of Zen at a fixed position nine bases from the 5′ end, the quencher is always within close proximity of the probe. As such, the initial background fluorescence signal is much lower. This makes subsequent changes easily detectable and functionally increases assay sensitivity. In addition, while traditional probes do not remain well quenched over 30 base pairs, the double-quenched probes maintain a consistently low background even at 40 base pairs or longer. Thus, double-quenched probes result in lower Cq values when compared to traditional probes. This leads to an increase in specificity without any loss in sensitivity or quenching efficiency. This is particularly useful for targets that are AT rich and require longer probes.