What do these terms mean to you?

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DNA sequencingDNA profiling/fingerprintingGene cloningTransformationMaking artificial genes

DNA Amplification

A crime scene(body tissue samples)

Fragments of DNA from a long extinct animal

A single viral particle (from an infection)

Using the technique called polymerase chain reaction (PCR), researchers are able to create vast quantities of DNA identical to trace samples. This process is also known as DNA amplification.

Many procedures in DNA technology require substantial amounts of DNA to work with, for example;

DNA sequencing

DNA profiling/fingerprinting

Gene cloning

Transformation

Making artificial genes

Samples from some sources,including those shown here,may be difficult to obtain inany quantity.

PCR EquipmentAmplification of DNA can be carried out with simple-to-use PCR machines called thermal cyclers (shown below).

Thermal cyclers are in common use in the biology departments of universities as well as other kinds of research and analytical laboratories.

The Process of PCR 1

Primer annealed

A DNA sample called the target DNA is obtained

DNA is denatured (DNA strands are separated) by heating the sample for 5 minutes at 98C

The temperature is lowered to 50C

Primers (short strands of mRNA) are annealed (bonded) to the DNA

The Process of PCR 2Nucleotides

Nucleotides

After one cycle, there are now two copies of the original sample.

The sample is heated to 60°C. A thermally stable Taq polymerase enzyme binds to the primers on each side of the exposed DNA strand.This enzyme synthesizes a complementary strand of DNA using free nucleotides.

Although only three cycles of replication are shown here, following cycles replicate DNA at an exponential rate and can make literally billions of copies in only a few hours.

Polymerase Chain ReactionPCR

cyclesNo. of target DNA strands

1 2

2 4

3 8

4 16

5 32

6 64

7 128

8 256

9 512

10 1024

11 2048

12 4096

13 8192

14 16 384

15 32 768

16 65 536

17 131 072

18 262 144

19 524 288

20 1 048 576

21 2 097 152

22 4 194 304

23 8 388 608

24 16 777 216

25 33 554 432

Cycle 1

Cycle 2

Cycle 3

Original DNASample

Can you match the definitions below to the key words on your worksheet?

• This enzyme can add complementary nucleotides to a DNA strand during DNA synthesis. It is similar to the human DNA polymerase responsible for copying your genome every time one of your body cells divides.

• These are short pieces of single-stranded DNA that match up to DNA sequences either side of the region of genomic DNA that you would like to copy. One is a forward primer and one is a reverse primer. When they have bound to the complementary sequences on the genomic DNA template strand, they show the Taq polymerase where to start DNA synthesis. The primers are responsible for making sure that only the region of interest is copied.

• This is double-stranded genomic DNA isolated from the cells of the organism being studied. It can be human DNA, plant DNA, mouse DNA, bacterial DNA, whatever DNA you would like to have copied!

• Free floating single nucleotides must be present in the reaction because they are what the Taq puts in place during DNA synthesis.

Lets act it out!

Man it´s hot in here!

98°C

Phew…I´m glad it´s cooler!

50°C

Seems to be warming up again!

60°C

Man it´s hot in here!

98°C

Phew…I´m glad it´s cooler!

50°C

Seems to be warming up again!

60°C

Now complete your PCR notes!

FILL IN THE BLANKS!!