TaqMan® (1, 2) is a process wherein a self-quenching probe is cleaved by the nuclease action of the polymerase. Figure 1 depicts an alternative to TaqMan®, and utilizes probes wherein fluorescent is due to fluorophore whose quenching is dependent on its stacking with other bases (3).

Figures 2 & 3 depict interesting apparently novel types of latently-fluorescent probes. These probes are designed to cycle upon a target sequence, such that there is a linear amplification of signal from this one single stranded target (for example that produced from RCA).

In Figure 2, the probe is similar to a TaqMan® probe except that it comprises one or more internucleotide bonds selectively cleaved upon double-stranded helix formation. Examples of such probes would be those which contain one or more ribonucleotides, which subsequent to hybridizing target, are cleaved by RNase H. Another example would be to form a mismatch or budge which is recognized and cleaved by a mismatch recognition enzyme (7, 8, 9). Notice the utility in sequence polymorphism analysis, as well as target sequence detection.

Notice that the process in Figure 2 could also be utilized with a probe as in Figure 1, wherein the fluorophore is quenched do to stacking. For example, a mismatch or budge comprising this fluorophore is activated upon cleavage by a mismatch or other repair enzyme.

Figure 3 shows yet another type of self-quenched cycling probe and its use. The essence of this process is to utilize a double-stranded selective exonuclease (4,5,6) to hydrolyze only target hybridized probes.

Notice the process could just as well have used probes comprising the pteridine nucleoside analogs described by Hawkin, et al (3).

Other ligand pairs whose signaling is modulated by proximity could also be utilized in the above examples.

This invention, as all the inventions described in this journal, is at the conceptual stage and it is hope that those with laboratories will investigate its full potential.

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