During PCR, what is the typical temperature used for the extension step with a DNA polymerase?

During PCR, what is the typical temperature used for the extension step with a DNA polymerase?

• A. 95°C
• B. 50-65°C
• C. 4°C
• D. 72°C

Answer: (D)

72°C is used during the extension step because standard thermostable DNA polymerases like Taq have their highest activity around this temperature. After the DNA is melted at 95°C and the primers have annealed around 50–65°C, the polymerase works best at about 72°C to efficiently add nucleotides to the 3' end of each primer, synthesizing the new strand. The exact extension temperature can vary with different polymerases, but 72°C is the classic optimum for the common enzymes used in PCR. The other temperatures correspond to the other phases: 95°C denatures the DNA, 50–65°C is for primer binding (annealing), and 4°C is a cold hold to pause or store the reaction.

72°C is used during the extension step because standard thermostable DNA polymerases like Taq have their highest activity around this temperature. After the DNA is melted at 95°C and the primers have annealed around 50–65°C, the polymerase works best at about 72°C to efficiently add nucleotides to the 3' end of each primer, synthesizing the new strand. The exact extension temperature can vary with different polymerases, but 72°C is the classic optimum for the common enzymes used in PCR. The other temperatures correspond to the other phases: 95°C denatures the DNA, 50–65°C is for primer binding (annealing), and 4°C is a cold hold to pause or store the reaction.