Lab 9 (PCR)

Question 1

4 components of PCR reaction

Answer 1

1) Template DNA containing the target sequence
2) DNA polymerase
3) dNTPs ( deoxynucleotide triphosphates )
4) 2 different primers

Question 2

Composition of the primers

Answer 2

synthetic oligonucleotides, about 17bp long

Question 3

DNA polymerase used

Answer 3

Taq polymerase from Thermus aquaticus

Question 4

Temperature that Taq polymerase resists

Answer 4

95-100 degrees

Question 5

Why is Taq polymerase ideal for PCR

Answer 5

It is heat resistant and fresh polymerase doesn’t have to be added following each round of DNA denaturation in thermal cycling of PCR

Question 6

What does heating the DNA does to it?

Answer 6

It denatures DNA template –> separates the 2 strands of DNA allowing the primers to anneal

Question 7

Name for the repeating heating and cooling process

Answer 7

Thermal cycling

Question 8

What is DNA sequencing?

Answer 8

in vitro DNA synthesis reaction but without amplification of the target sequence.

Question 9

What are the differences between DNA sequencing and PCR? (3)

Answer 9

1) DNA sequencing uses 1 primer only
2) DNA sequencing has dideoxynucleotides (terminators)
3) DNA sequencing results in a mixture of newly-synthesize fragments, each terminating at a different nucleotide location.