In PCR, why is the mixture cooled to 40°C?

• A. Denature DNA
• B. Polymerize nucleotides
• C. Bind primers to DNA
• D. Activate Taq polymerase

Answer: (C)

Primer annealing is what happens when the reaction is cooled to a temperature like 40°C. After the DNA strands have been separated, the short primers need to find and bind to their complementary sequences on the single‑stranded DNA. This binding relies on the temperature being low enough for stable base pairing but not so low that primers bind nonspecifically. Once the primers are bound, the next step raises the temperature so the DNA polymerase can extend from the 3′ end of each primer. That’s why 40°C is used here—to allow primers to bind to their targets. Denaturation occurs at a much higher temperature, and polymerization happens during the extension step, not during annealing. Activating Taq polymerase is not the purpose of this cooling step.