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Objectives
You should be able to describe the biological principles
underlying each of the following:
 The use of the polymerase chain reaction (PCR) to
make large amounts of DNA from very small samples
 The use of genetic fingerprinting
Criminals – Watch Out!
 Scientists can now identify criminals from the smallest
bit of DNA.
 Wherever you go, you leave a trail of DNA behind.
 We consistently shed hairs and flakes of skin.
 It is in these hair and skin cells that DNA is found!
 You DNA is your GENETIC FINGERPRINT!
Polymerase Chain Reaction(PCR)
 PCR is an method by which DNA can be replicated in
the lab.
 It can be used to create millions of copies of DNA in
just a few hours.
 It is essential in forensic science as very small samples
of DNA are difficult to analyse.
 This process amplifies DNA, so that it can be analysed.
What do you need?
1.
RNA primers provide the starting sequence for DNA
replication. They also stop the two DNA strands
from joining together.
2.
DNA nucleotides containing the bases adenine,
guanine, cytosine and thymine.
3.
Enzyme DNA polymerase.
The Stages of PCR
Strand Separation
DNA heated at 95°C
for 5mins
Mix with Primers
(RNA strands)
C
Binding of Primers
Mixture cooled to
40°C
Mix with
Free Nucleotides
DNA Polymerase
REPEAT
CYCLING
With every cycle the amount
of DNA doubles
DNA Synthesis
Mixture heated to
70°C
(optimum temp. for
DNA polymerase)
The Double Stranded DNA Molecule
A
A G G
T
C
A
C
T
T
T
T
A G
T
G A
A
C C
Heat to 950C to separate the DNA strands
DNA Strand
A
A G G
C
T
T
C
T
A
C
T
C C
T
RNA Primers
C
T
T
A G
T
T
T
G A
A
DNA Strand
Cool to 400C to allow primers to bind (anneal) to DNA
DNA Polymerase
G
Original DNA strand
A
A G G
T
C
A
T
T
A G
T
Primer
G
C C
T
G
T
C
C
Nucleotides join on
Free DNA nucleotides
A
A
T
G
C
T
G
G
C C
Nucleotides join on
T
C
A G
A
T
C
T
Primer
G A
G
T
A
Free DNA
nucleotides
Original DNA strand
Mix with DNA polymerase and free nucleotides and heat to 700C
Advantages
 It is a very rapid process
 Does not require living cells
 Is useful when we want to introduce a gene into
another organism
Disadvantages
These are also the advantages of In vivo cloning:
 There is risk of contamination by unwanted DNA
 It cannot cut out specific genes
 Does not produce transformed bacteria
Summary Questions
 In the polymerase chain reaction, what are the
‘primers’?
 What is the role of these primers?
 Why are two different primers required?
 When DNA strands are separated in the PCR, what
type of bond is broken?
 It is important in the PCR that the fragments of DNA
used are not contaminated with any other biological
material. Suggest a reason why.