CSMLS Molecular Diagnostics

Question 1

What is the primary purpose of the denaturation step in PCR?

Accepted Answer

To separate double-stranded DNA into single strands

Answer

Denaturation at 94–96°C breaks the hydrogen bonds between complementary base pairs, separating the double-stranded DNA into single-stranded templates for subsequent primer annealing.

Question 2

Why is Taq polymerase used in PCR instead of a standard DNA polymerase?

Accepted Answer

It is thermostable and survives repeated heating to 94°C

Answer

Taq polymerase, derived from Thermus aquaticus, is thermostable and retains activity through the repeated high-temperature denaturation cycles required in PCR.

Question 3

In quantitative real-time PCR (qPCR), what does the Ct (cycle threshold) value represent?

Accepted Answer

The cycle at which fluorescence crosses the detection threshold

Answer

The Ct value is the PCR cycle number at which the fluorescence signal rises above background threshold; a lower Ct indicates a higher initial amount of target nucleic acid.

Question 4

Which technique is used to detect RNA viruses such as HIV or Hepatitis C virus by molecular methods?

Accepted Answer

Reverse transcriptase PCR (RT-PCR)

Answer

RT-PCR uses reverse transcriptase to first convert the viral RNA into complementary DNA (cDNA), which is then amplified by standard PCR to enable detection of RNA-based targets.

Question 5

SYBR Green dye in qPCR generates signal by:

Accepted Answer

Intercalating into any double-stranded DNA formed during amplification

Answer

SYBR Green intercalates non-specifically into any double-stranded DNA, producing fluorescence proportional to the amount of double-stranded PCR product accumulated.

CSMLS - Canadian Society for Medical Laboratory Science Practice Test

CSMLS - Canadian Society for Medical Laboratory Science Practice Test

CSMLS Molecular Diagnostics