DNA Profiling

DNA profiling is a forensic technique whereby scientists compare similarities between two DNA samples. The DNA fragments are amplified by PCR then separated by electrophoresis to create a distinctive banding pattern. It has very important applications - from determining paternity, identifying a new species or implicating a suspect in a crime.

 
 

DNA Profiling

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DNA profiling is a technique which can be used to analyse a sample of DNA (e.g. one found at a crime scene) and compare it to DNA samples taken from the suspects. The DNA sample will be collected (this is usually blood, saliva or semen) and amplified using PCR. The PCR products are separated using gel electrophoresis, which separates the DNA fragments according to length. The gel is visualised using UV light and the banding patterns from the suspect’s DNA can be compared with that found at the crime scene. The same technique can also be used to identify genetic relationships between people (as in paternity testing) or to determine evolutionary relationships between organisms.

Polymerase Chain Reaction (PCR)

PCR is a technique used to amplify fragments of DNA. This is important because the DNA sample collected at a crime scene doesn’t contain enough material for accurate analysis. Before PCR is carried out, you need to prepare a mixture containing the following:

  • DNA sample

  • Free DNA nucleotides

  • Primers (these are short pieces of DNA which bind to the beginning of the DNA fragment and initiate replication)

  • The enzyme DNA polymerase which has been extracted from thermophilic (heat-loving) bacteria.

PCR is then carried out in the following stages:

  1. Separation of the DNA strands - the mixture is heated to 95oC which causes hydrogen bonds between the DNA strands to break.
  2. Annealing of the primer - the mixture is then cooled to around 60oC which allows the primer to anneal to the DNA.
  3. DNA synthesis - The temperature is increased to 72oC which is the optimum temperature for DNA polymerase. DNA polymerase forms a new DNA strand from catalysing the formation of phosphodiester bonds between the free DNA nucleotides which align along the DNA template strand by complementary base pairing rules.
 
 

Around 30-40 cycles of PCR are carried out, which generates millions of DNA fragments. Each PCR cycle doubles the amount of DNA, so huge numbers of DNA fragments can be quickly generated.

DNA stained with ethidium bromide is separated on by gel electrophoresis. Image: School of Natural Resources from Ann Arbor – CC-BY 2.0

DNA stained with ethidium bromide is separated on by gel electrophoresis. Image: School of Natural Resources from Ann Arbor – CC-BY 2.0

Fluorescent tags

So that the DNA can be visualised, we add a fluorescent molecule which binds to the DNA and makes it visible when exposed to UV light. A common fluorescent tag is ethidium bromide, which inserts itself between the DNA bases and gives off fluorescence under UV light.

Electrophoresis

Once the DNA fragments have been amplified and stained with a fluorescent dye, they need to be separated. This is done using gel electrophoresis, which separates the DNA strands according to length. It works because DNA is negatively charged, which means it will move towards a positive charge when placed in an electric field. Shorter DNA fragments travel through the gel more quickly, which means they will travel a longer distance than the larger DNA fragments. Gel electrophoresis is carried out in the following steps:

  • An agarose gel is prepared which contains a row of wells at the top of the gel. The gel is placed into a tank containing buffer solution which is able to conduct electricity.

  • The DNA sample is mixed with a loading dye - this turns the DNA mixture a dark colour and helps you see what you’re doing. A fixed volume of the DNA samples are pipetted into the wells.

  • An electrical current is passed through the gel and the DNA will begin to move towards the bottom of the gel (towards the anode).

  • Once the dye has reached the bottom, the electricity is turned off and the banding pattern is visualised under UV light.

 
 
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Did you know..

In 2003, a woman called Lydia Fairchild was accused of a scam when a DNA test revealed that she was not the mother of her own children. At risk of having her kids taken from her, it was eventually discovered that Lydia is a chimera - she contains DNA from two origins. Before she was born, cells from her unborn twin sister merged with her own which meant that the cells of her blood contained different DNA to those found in other parts of her body.