What is the general workflow of RT-PCR when detecting RNA viruses?

What is the general workflow of RT-PCR when detecting RNA viruses?

• A. RT-PCR uses a thermostable RNA polymerase to amplify RNA directly.
• B. Reverse transcription converts RNA to cDNA, then PCR amplifies target DNA; used for detection/quantification of RNA viruses.
• C. RT-PCR removes RNA and leaves it unamplified.
• D. RT-PCR relies on DNA polymerase to convert RNA into RNA.

Answer: (B)

The central idea is that RNA viruses are detected by turning their RNA into DNA first, because PCR works on DNA. In RT-PCR, an enzyme called reverse transcriptase copies the viral RNA into complementary DNA (cDNA). Once this DNA template is available, standard DNA polymerase enzymes perform PCR to amplify the target cDNA. This makes it possible to detect and often measure how much viral RNA was present in the sample. One-step and two-step formats exist: one-step combines reverse transcription and PCR in the same tube, while two-step does the reverse transcription first to make cDNA, then uses that cDNA in a separate PCR reaction. The readout, especially in real-time RT-PCR, tracks the amplified product to quantify the amount of viral RNA initially present. Why the other ideas don’t fit: PCR can’t start from RNA because DNA polymerases require a DNA template. Directly amplifying RNA would be incorrect. Leaving RNA unamplified wouldn’t produce the required DNA template for amplification. And converting RNA to RNA isn’t the typical mechanism here; it’s RNA to DNA via reverse transcription, followed by DNA amplification.

The central idea is that RNA viruses are detected by turning their RNA into DNA first, because PCR works on DNA. In RT-PCR, an enzyme called reverse transcriptase copies the viral RNA into complementary DNA (cDNA). Once this DNA template is available, standard DNA polymerase enzymes perform PCR to amplify the target cDNA. This makes it possible to detect and often measure how much viral RNA was present in the sample.

One-step and two-step formats exist: one-step combines reverse transcription and PCR in the same tube, while two-step does the reverse transcription first to make cDNA, then uses that cDNA in a separate PCR reaction. The readout, especially in real-time RT-PCR, tracks the amplified product to quantify the amount of viral RNA initially present.

Why the other ideas don’t fit: PCR can’t start from RNA because DNA polymerases require a DNA template. Directly amplifying RNA would be incorrect. Leaving RNA unamplified wouldn’t produce the required DNA template for amplification. And converting RNA to RNA isn’t the typical mechanism here; it’s RNA to DNA via reverse transcription, followed by DNA amplification.