Teacher notes: The polymerase chain reaction
What is the lesson about?
This resource can be used as a virtual teaching tool to show the stages and
amplification of a DNA section by the polymerase chain reaction (PCR).
This technique is widely used in molecular biology to amplify a sequence of
DNA and has a range of applications, including DNA cloning, medical
diagnosis of, for example, hereditary disease, as well as forensic science
(genetic fingerprinting).
The method was created in 1983 by Kary Mulli s, who later won the Nobel
Prize (1993) for chemistry for his work.
Movies can be controlled by clicking forward and back arrow keys.
Learning outcomes
Higher Biology (revised), Unit 1.
1. The structure and replication of DNA b) (ii) Polymerase chain reaction
Aims
Students will be able to describe the PCR process as a technique that relies on
primers, enzymes and a series of heating and cooling cycles to amplify a
specific region of DNA.
The polymerase chain reaction – title slide
In laboratories, PCR is automated within a machine called a thermal cycler.
This controls the temperature changes required in the experiment
automatically. Small tubes containing the reaction mixture (DNA, primers,
DNA polymerase, buffer totalling typically 100 µl) are placed in the machine.
The duration of each interval and exact temperature will be optimised by
scientists for their specific reaction.
(Click) Slide 1: Schematic diagram of double stranded DNA
(Click) to highlight thermometer.
(Click) to highlight DNA sequence to be amplified. All the chemicals
required for the PCR reaction are added at the start. The temperature controls
the progress of the reaction.
TEACHER’S NOTES: THE POLYMERASE CHAIN REACTION (H, BIOLOGY)
© Learning and Teaching Scotland 2011
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(Click) Denaturation. Temperature rises to typically 94°C. This breaks the
hydrogen bonds between bases, strands separate to generate single-stranded
DNA.
(Click) Primer annealing. The temperature drops to typically 58°C. This
allows pre-made primers (complementary to specific target sequences at the
two ends of the region to be amplified) to bind.
(Click) Elongation. The temperature rises to typically 72°C, activating the
DNA polymerase enzyme, which then reads the sequence of the DNA, adding
complementary bases. Most PCR reactions use Taq polymerase, an enzyme
isolated from Thermus aquaticus. This bacterium is found living in hot
oceanic thermal vents and so contains the heat-stable polymerase. The
polymerase assembles the complementary strand. This creates two new
double-stranded DNA molecules.
For the next cycles ask student to predict the different stages, ie when the
temperature first rises what will happen? ( Denaturation to two single strands.)
(Click) The process repeats, with the temperature rising to 94°C to produce
four single-stranded DNA molecules.
(Click) The temperature drops to 58°C and primers anneal.
(Click) The temperature rises to 72°C and the DNA polymerase reads the
sequence, creating four double stands.
(Click) The cycle repeats, temperature rises – denaturation.
(Click) Primers anneal.
(Click) Elongation to create eight double strands.
An average PCR reaction will include 25–35 cycles, running for 90 min. The
DNA amplification is exponential, resulting in a large amplification of the
initial sample.
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TEACHER’S NOTES: THE POLYMERASE CHAIN REACTION (H, BIOLOGY)
© Learning and Teaching Scotland 2011
Throughout the lesson you may want to prompt students by asking them to
describe what is happening in the animation as cycles repeat. Written answer s
may also be sought:
1.
2.
The PCR process requires heating and cooling of the DNA reaction
mixture. Describe what process is occurring in the reaction mixture at
the following temperatures: (Teacher answers in italics)
94°C:
Denaturation. Temperature rises to typically 94 °C. This breaks
the hydrogen bonds between bases, strands separate to
generate single-stranded DNA.
58°C:
Primer annealing. The temperature drops to typically 58 °C.
This allows pre-made primers – complementary to specific
target sequences at the two ends of the region to be amplified –
to bind.
72°C:
Elongation. The temperature rises to typically 72°C, activating
the DNA polymerase enzyme, which then reads the sequence of
the DNA, adding complementary bases. Most PCR reactions
use the Taq polymerase, an enzyme isolated from Thermus
aquaticus. This bacterium is found living in hot oceanic
thermal vents and so has a heat-stable polymerase. The
polymerase assembles the complementary strand. This creates
two new double-stranded DNA molecules.
Research some applications of the PCR process. (Starting points may
include DNA cloning, medical diagnosis of , for example, hereditary
disease, genetic fingerprinting in forensic science .)
TEACHER’S NOTES: THE POLYMERASE CHAIN REACTION (H, BIOLOGY)
© Learning and Teaching Scotland 2011
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