Chapter 4: 4.8 In Vitro DNA Replication: Polymerase Chain Reaction (PCR)

Question 1

What does “in vitro method of replication/amplifying DNA” mean?

Answer 1

Uses the same concepts of DNA replication to amplify a region of choice from a target DNA in a tube

Question 2

What does PCR stand for?

Answer 2

Polymerase Chain Reaction

Question 3

What does a test tube for PCR contain?

Answer 3

• A thermostable DNA polymerase (Taq polymerase)
• Deoxyribonucleotides of all varieties
• The DNA template to be amplified
• Primers for both strands of the DNA

Question 4

What happens to carry out the process?

Answer 4

A thermocycler
* Cycles through various temperatures to carry out the process

Question 5

What are the 3 steps involved in PCR?

Answer 5

1. Denaturation
2. Annealing
3. Elongation

Question 6

Differences from DNA Replication in the Cell:

Explain the difference in unwinding of DNA

Answer 6

1. Uses heat to unwind DNA strands instead of helicase
2. Completely denatures DNA into single strands (not a progressive unwinding)

Question 7

Differences between PCR and DNA Replication in the Cell:

Explain the difference in primers

Answer 7

Primers are artificially designed and already provided in the test tube

Question 8

True or False:

PCR primers are made of RNA, like the cell

Answer 8

False, they are made of DNA

Question 9

What do artificially created primers in PCR allow us to do?

Answer 9

Primers dictate where the replication reaction will occur
* Allows us to specify the region of interest

Question 10

Differences from DNA Replication in the Cell:

Explain the differences in length of DNA replicated

Answer 10

Only a small region is amplified as opposed to the whole DNA template

Question 11

Differences from DNA Replication in the Cell:

Explain the differences in the number of copies of DNA made

Answer 11

Millions of copies of the target region is made as opposed to only two copies that occur in the cell before it divides

Question 12

Differences from DNA Replication in the Cell:

Explain the differences in lagging and leading strand

Answer 12

No lagging or leading strands involved

Question 13

In PCR:

What are the requirements of the primer design?

Answer 13

• Primers must flank the region of DNA that we want to amplify and be complementary to the strands
• They are usually 18-22 base pairs