In a TaqMan real-time PCR assay, fluorescence signal is generated when:
- A The intercalating dye binds any double-stranded DNA nonspecifically
- B The probe melts off the template at the annealing temperature, releasing free dye
- C Taq polymerase cleaves the hybridized probe via its 5' nuclease activity, separating reporter from quencher ✓
- D The reverse transcriptase incorporates labeled nucleotides into cDNA
Correct answer: C. Taq polymerase cleaves the hybridized probe via its 5' nuclease activity, separating reporter from quencher
Explanation
The TaqMan probe carries a reporter fluorophore at its 5' end and a quencher at its 3' end. While intact, the quencher suppresses reporter emission. When Taq polymerase extends the primer and reaches the probe, its 5' exonuclease activity cleaves the probe, physically separating reporter from quencher and releasing fluorescence proportional to amplicon accumulation. SYBR Green, not TaqMan, works by nonspecific binding to any double-stranded product.
Reference: Koneman's Color Atlas and Textbook of Diagnostic Microbiology, 7th ed.
High-yield for: NEET PGINI-CETNExTFMGEUSMLEPLABMRCP
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