Practice Test GeeksPCR - Polymerase Chain Reaction Certification Practice Test
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Principles & Techniques Flashcards

7 cards from real PCR practice questions. Tap to flip, then mark Knew It or Still Learning — missed cards come back until you master them.

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  1. In real-time qPCR, what does the cycle threshold (Ct) represent?

    Answer: The cycle at which fluorescence crosses a set threshold above background

    Ct is the cycle where signal first rises detectably above background, and it is inversely related to starting template amount.

  2. A qPCR standard curve has a slope of -3.32. What amplification efficiency does this indicate?

    Answer: About 100%

    Efficiency = 10^(-1/slope) - 1, and a slope of -3.32 gives about 100%.

  3. How does a TaqMan hydrolysis probe generate fluorescence?

    Answer: The polymerase's 5'→3' exonuclease cleaves the probe, separating reporter from quencher

    Cleaving the bound probe releases the reporter dye from the quencher, so fluorescence rises.

  4. What is the main drawback of SYBR Green chemistry compared with probe-based qPCR?

    Answer: It binds any double-stranded DNA, including primer-dimers and nonspecific products

    SYBR Green is not sequence-specific, so any dsDNA adds to the signal.

  5. After a SYBR Green qPCR run, a melt curve shows two distinct peaks. What does this most likely indicate?

    Answer: More than one product, such as a primer-dimer alongside the target

    Each distinct melting peak reflects a dsDNA species with a different Tm.

  6. In the ΔΔCt method, a target gene shows a ΔΔCt of -2. What is its fold change relative to the calibrator, assuming 100% efficiency?

    Answer: 4-fold increase

    Fold change = 2^-ΔΔCt = 2^2 = 4.

  7. What is the purpose of a passive reference dye such as ROX in qPCR?

    Answer: To normalize for well-to-well differences in fluorescence unrelated to amplification

    ROX corrects for pipetting volume differences and optical variation across wells.

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