Locating a desired DNA sequence
A DNA probe is a short single-stranded section of DNA that is complementary to the section of DNA being investigated. The probe is labelled in one of two ways:
- DNA samples are treated with restriction enzymes to cut them into fragments
- The DNA samples are placed into wells cut into the negative electrode end of an agarose gel
- A DNA standard is added to one well – the fragments are of known length so can be used to estimate the size of the fragments in the samples
- The gel is immersed in a tank of buffer solution and an electric current is passed through the solution for around 2 hours
- DNA is negatively charged because of its phosphate groups so is attracted to the positive electrode
- Shorter lengths of DNA move faster than longer lengths so move further in the time the current is run
- The position of the fragments can be shown by using a dye that stains DNA molecules (see above)
You could try carrrying out this procedure in a virtual lab by clicking here!
Amplifying a DNA sequence
PCR stands for the polymerase chain reaction. It is used to make lots of copies of DNA and is even used by the police to give a large enough quantity of DNA for testing from the smallest blood or other sample.
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The DNA sample is mixed with a supply of DNA nucleotides and DNA polymerase
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The mixture is heated to 95°C. This breaks the hydrogen bonds holding the strands together, so the samples are now single stranded
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Primers (short lengths of single stranded DNA) are added
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The temperature is reduced to 55°C to allow the primers to bind and form small sections of double stranded sections;
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DNA polymerase can bind to these double-stranded sections;
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The temperature is raised to 72°C. The enzyme extends the double stranded section by adding nucleotides to the unwound DNA;
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When the DNA polymerase reaches the other end of the DNA, a new, double stranded DNA molecule is generated;
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The whole process can be repeated many times so the amount of DNA increases exponentially.
You can watch this procedure being carried out by clicking here and going to ‘techniques – amplifying’ or you could carry out the procedure for yourself in a virtual lab here!