Question

1. 95°c for 2 minutes
2. 95°c for 30 seconds
3. 58.1°c for 30 seconds
4. 72°c for 45 seconds
5. repeat steps 2 - 4 thirty (30) times
6. 72°c for 3 minutes
7. hold at 16°c indefinitely

which steps represent the denaturation, annealing and extension steps?
○ a. step 2 = denature; step 3 = anneal; step 4 = extend
○ b. step 4 = denature; step 6 = anneal; step 7 = extend
○ c. step 2 = denature; step 4 = anneal; step 6 = extend
○ d. step 3 = denature; step 4 = anneal; step 6 = extend
clear my choice

Explanation:

• Denaturation: In PCR (Polymerase Chain Reaction), denaturation occurs at high temperatures (around \(95^{\circ}C\)) to separate the double - stranded DNA into single strands. Step 2 (\(95^{\circ}C\) for 30 seconds) is a denaturation step.
• Annealing: Annealing is the process where primers bind to the single - stranded DNA. It occurs at a lower temperature (usually \(50 - 65^{\circ}C\)). Step 3 (\(58.1^{\circ}C\) for 30 seconds) is an annealing step.
• Extension: Extension is when the DNA polymerase synthesizes new DNA strands. It occurs at a temperature (\(70 - 75^{\circ}C\)) where the DNA polymerase is most active. Step 4 (\(72^{\circ}C\) for 45 seconds) is an extension step.

Answer:

A. Step 2 = Denature; Step 3 = Anneal; Step 4 = Extend