Minor U12-type with GU-AG introns (hybrid type)

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#Id: 2551

#Unit 13. Methods in Biology

5′ nuclease assay (TaqMan assay). PCR is undertaken with a fluorescence reporter/quencher pair (called RQ probe). 

TLS Online TPP Program

#Id: 2552

#Unit 13. Methods in Biology

During extension by Taq polymerase, the probe is cleaved owing to the nuclease activity of the polymerase.

TLS Online TPP Program

#Id: 2553

#Unit 13. Methods in Biology

Due to release of the reporter, there is a detectable increase in fluorescence, which is monitored in real-time PCR experiments.

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#Id: 2587

#Unit 13. Methods in Biology

In case of nested PCR, the target sequence is amplified using a pair of PCR primers, and a portion of the amplification product is reamplified using another pair of PCR primers complementary to the regions located immediately beyond the 3'-ends of the first pair of primers.

TLS Online TPP Program

#Id: 2588

#Unit 13. Methods in Biology

In case of nested PCR, the inner primers have a lower annealing temperature than that of the outer pair. For the initial several cycles of PCR the higher denaturation and annealing temperatures are used; as a result, only the outer pair of primer is able to prime amplification. 

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#Id: 2589

#Unit 13. Methods in Biology

Nested PCR follows the 'drop in drop out' PCR phenomenon.