What is the correct order of steps in a PCR cycle?

• A. Denaturation, annealing, extension.
• B. Denaturation, extension, annealing.
• C. Annealing, Denaturation, Extension.
• D. Extension, Denaturation, Annealing.

Answer: (A)

PCR cycles rely on three steps in a precise order: denaturation, annealing, extension. First, denaturation uses heat to separate the double-stranded DNA into single strands, giving the template accessible for primer binding. Next, annealing cools the mixture so short DNA primers can bind to their complementary sequences on the single-stranded templates. Finally, extension uses DNA polymerase to add nucleotides to the primers, synthesizing new DNA and producing double-stranded products again. This order is essential because primers cannot bind to double-stranded DNA, and polymerase needs bound primers to begin synthesis. If you tried to extend before primers are bound, or to anneal before the strands are separated, amplification wouldn’t occur efficiently. So the correct sequence is denaturation, annealing, extension.