Pcr Practical 1

Question 1

Which transformants were put into PCR for analysis

Answer 1

Pblt101 (to see if the insert was there)

Question 2

What is the first step of PCR in a thermocycler

Answer 2

Denaturation at 94C (disrupts h bonds)

Question 3

What happens after densturation

Answer 3

Set to 55C to allow primers to anneal

Question 4

What were the primers used in tube 1,2,3

Answer 4

1- forward universal primer and reverse universal primer (included lacz and insert)

2- FUP and internal primer (195bp)

3- IP and RUP (440bp)

Question 5

What happens after primers set

Answer 5

Extension/elongation

Taq polymerase binds from the 3’ oh

Adds DNTPs

Question 6

Which cycle do double stranded copies become the correct length (short)

Answer 6

3rd cycle

Question 7

What are the 4 issues with PCR products

Answer 7

Not every dna will denature

Taq might not copy whole amplicon

Primers/taq may not binds

Components might deplete

Question 8

Which organism does taq come from

Answer 8

Thermus aquaticus