PCR - L1

Question 1

Why do we use PCR?

Answer 1

sensitive, cheap, specific, rapid and robust

Question 2

What are the elements present in a PCR tube

Answer 2

Template, 2 primers, polymerase, dNTP, Mg, buffer

Question 3

Describe the characteristics of primers

Answer 3

small ss DNA, 6-30 bases, chemically synthesised

Question 4

State the role of Mg in PCR

Answer 4

Cofactor for enzymes, enhances enzymic activity enabling catalysis

Question 5

What is the ideal length for primers

Answer 5

18-24

Question 6

What are primers mainly composed of

Answer 6

40-60% GC

Question 7

What do primers start and end with

Answer 7

1-2 GC pairs

Question 8

What is the melting temp

Answer 8

50-60

Question 9

What is the range of temp that primers should have between each other

Answer 9

5

Question 10

What end must be complimentary to the DNA

Answer 10

3’

Question 11

Should primer pairs have complimentary regions

Answer 11

NO

Question 12

In what form is Mg used

Answer 12

MgCl2

Question 13

What is the optimal buffer pH

Answer 13

8-9.5

Question 14

What are the components of buffer

Answer 14

Tris HCL and KCl (or ammonium sulphate)

Question 15

What are the three stages of PCr

Answer 15

denaturation, annealing, elongation

Question 16

How many cycles in PCR

Answer 16

30

Question 17

is a different primer used in the second cycle?

Answer 17

yes

Question 18

What temp does denaturing occur

Answer 18

95

Question 19

WHat temp does anneal occue

Answer 19

55

Question 20

What temp does elongation occur

Answer 20

72

Question 20

هلا

Answer 20

اهلين

Question 21

What is the end product of PCR (how much DNA)

Answer 21

1 billion copies

Question 22

What is the PCR marker and what colour is it

Answer 22

Nancy520 (red)

Question 23

What type of gel do we use to detect PCR products

Answer 23

agarose gel