The 5‵Nuclease Assay
In the 5‵ nuclease assay, an oligonucleotide called a TaqMan® Probe is added to the PCR reagent master mix. The probe is designed to anneal to a specific sequence of template between the forward and reverse primers.
The probe sits in the path of the enzyme as it starts to copy DNA or cDNA. When the enzyme reaches the annealed probe the 5 exonuclease activity of the enzyme cleaves the probe, Figure 12 through 14.
Figure 12: The 5‵Nuclease Assay
Figure 13: Polymerase collides with TaqMan® Probe
Figure 14: Cleavage of the TaqMan® Probe
