Question 3. This question is meant for you to think about what would happen in a

Question

Question 3. This question is meant for you to think about what would happen in a PCR reaction if you only added one primer, rather than using both a forward and reverse primer. Consider the following DNA molecule, used as the only DNA template molecule to initiate a PCR reaction 5'-TTTTTTTTTTTTTTTTTTTTTCACACA-3' 3' -AAAAAAAAAAAAAAAAAAAAAGTGTGT-5 Now consider adding a single primer that has the sequence: 5' -TGTGTG-3' A) Which template strand would be used in the reaction? (top or bottom?) B) What would be the sequence of the DNA that is synthesized in the reaction? C) Could the DNA that is being synthesized in one cycle (the DNA sequence from B) be used as a template for the next cycle of the reaction? What effect does this have on the ability of PCR to amplify the target sequence?

Explanation / Answer

a) Top as the primer is complementary to the top.

b) 5' TGTGTGAAAAAAAAAAAAAAAAAAAAA 3' (based on complementarity)

C) No, since the primer is only for the top strand which gives complementary sequence after one cycle. This new strand is not complementary with the primer. Hence in every cycle, the primer will have to bind to the parent strand itself. This will impede the purpose of PCR as no chain reaction and thereby exponential amplification will take place.

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