International A Level Biology / Decomposition and forensics
Gel electrophoresis revision notes
Edexcel International A Level Biology (YBI11), chapter 19: Decomposition and forensics, specification point 6.18. Free lesson.
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Gel electrophoresis sorts DNA fragments by length. It is used to compare DNA samples and estimate fragment sizes by seeing how far the fragments travel through a gel.
How restriction enzymes make fragments
Before DNA is separated, a restriction enzyme cuts it at a specific short base sequence called a recognition site. DNA length is measured in base pairs (bp). The distances between the ends of the molecule and the cut sites give the lengths of the fragments. The same restriction enzyme cuts the same DNA at the same sites, so it makes the same fragment lengths each time.
How to run a gel
Agarose, a jelly made from seaweed, forms a slab with small pockets called wells at one end. The gel sits in a tank and is covered with buffer solution, which conducts the current and keeps the pH steady. Follow the steps in order when describing the method.
Why the fragments separate by length
DNA has a negative charge because its sugar-phosphate backbone contains negatively charged phosphate groups. When the current is switched on, the fragments move towards the anode, the positive electrode. The gel is a mesh with tiny pores. Short fragments squeeze through more easily, so they travel further in the same time; longer fragments are held back near the wells. The fragments move because the current pulls them towards the anode, not because of diffusion down a concentration gradient. The wells must be at the cathode. If they are at the anode, the DNA runs straight out of the gel.
How to read the bands
All the fragments move away from the wells in the same direction, towards the anode. Longer fragments form bands nearer the wells, while shorter fragments travel further and form bands nearer the anode. Compare a sample band with the DNA ladder to estimate its length.
How the bands are made visible
DNA is colourless, so you need to make it visible after it has been separated. A stain binds to DNA and shows every band. A probe is labelled DNA that pairs with a matching sequence, so it only shows fragments containing its complementary sequence. A radioactive probe produces visible bands on X-ray film. A fluorescent probe glows under ultraviolet light.