Fig. 2. Real-time PCR using Taqman probes. (A) A schematic of PCR in the presence of a Taqman probe. (1) Probes and primers anneal to target sequence. Taqman probes have two covalently linked fluorescent dyes: a reporter (R) and a quencher (Q). On the probe, the reporter dye emission is quenched. (2) During each extension cycle, the 5′→3′ exonuclease activity of Taq DNA polymerase cleaves the reporter dye from the probe. (3) Once separated from the quencher, the reporter dye emits its characteristic fluorescence, which is measured in every cycle by the ABI Prism 7700 sequence detector. (B) A representative panel of data generated by the ABI Prism 7700 sequence detector software for Taqman probe 5.613 on samples immunoprecipitated from 6C2 nuclei. IP indicates the signal for the fluorescent reporter detected in each cycle for the immunopreciptated target sequence. Ref indicates a similar analysis of the target sequence abundance in total input DNA. Ct indicates the cycle threshold number. Below the data panel is a sample calculation to determine the fold difference between the IP sample and the reference sample.