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Abstract

Disclosed are compositions and methods for real-time detection of rolling circle amplification products. Real-time detection is detection that takes place during the amplification reaction or operation. Real-time detection can be accomplished by, for example, using fluorescent change probes and/or primers during amplification. The fluorescent signals can be proportional to the amount of amplification product. The amplification can be multiply-primed rolling circle amplification in which replication of a circular template is primed at a plurality of sites on the circular template. Multiply-primed RCA increases the sensitivity of singly-primed rolling circle amplification. Multiply-primed RCA can be performed using a single primer (which hybridizes to multiple sites on the amplification target circle) or multiple primers (each of which can hybridize to a single site on the amplification target circle or multiple sites on the amplification target circle). Fluorescent change probes and primers are probes and primers that involve a change in fluorescence intensity or wavelength based on a change in the form or conformation of the probe or primer and nucleic acid to be detected, assayed or replicated.

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First Claim