What are the three major steps in PCR Write them in order of

What are the three major steps in PCR? Write them in order of occurrence? Denaturation, primer annealing, and extension What is the purpose of the number of cycles during PCR? What happens if not enough cycles are used? What happens if too many cycles are used? At what temperature is denaturation performed? 95C At what temperature is primer annealing performed? 45-55C At what temperature is elongation performed? 70-72C Which of the three steps has the most temp variation? Explain why this is What is the relationship between fragment length and time during PCR elongation? What does RT-PCR stand for? Reverse transcriptase PCR What enzyme is essential for RT-PCR? Reverse transcriptase What is the purpose of or product from RT-PCR? To what is RT-PCR an alternative? qPCR

Solution

6)

Polymerase chain reaction or PCR is useful to amplify small sample of DNA producing multiple copies.

7)

PCR is used to amplify the DNA copies. Each cycle produces 2 copies of DNA. If enough cycles are not used in PCR, the amount of DNA required would be reduced.

8)

If the number of cycles exceed in PCR, then the amount of DNA amplified is more than the required amount.

12)

Most temperature variation is observed in denaturation and primer annealing steps. This because of heating and cooling alternated in these steps. In elongation step of PCR, the Taque DNA polymerase obtained from a thermophile Thermus aquaticus is used.

13)

During PCR elongation, the DNA polymerase used and fragment length influence the time duration of the process. The efficiency of DNA polymerase may conclude the process at rapid rate. More is the fragment length and more is the time taken to complete the process.