The distance the DNA has migrated in the gel can be judged visually by monitoring the migration of the loading buffer dye.Shorter lengths of DNA move faster than longer lengths so move further in the time the current is run.The electrical current is then turned on so that the negatively charged DNA moves through the gel towards the positive side of the gel.When this is done the lid is placed on the electrophoresis tank making sure that the orientation of the gel and positive and negative electrodes is correct (we want the DNA to migrate across the gel to the positive end).The prepared DNA samples are then pipetted into the remaining wells of the gel.The fragments in the marker are of a known length so can be used to help approximate the size of the fragments in the samples. A DNA marker (also known as a size standard or a DNA ladder) is loaded into the first well of the gel.A dye is added to the sample of DNA prior to electrophoresis to increase the viscosity of the sample which will prevent it from floating out of the wells and so that the migration of the sample through the gel can be seen.The type of buffer used depends on the approximate size of the DNA fragments in the sample. The buffer conducts the electric current. The gel is then placed into an electrophoresis tank and electrophoresis buffer is poured into the tank until the surface of the gel is covered.Once the gel has cooled and solidified (it will now be opaque rather than clear) the comb is removed.The molten gel is then poured into a gel casting tray and a “comb” is placed at one end to make wells for the sample to be pipetted into.To make a gel, agarose powder is mixed with an electrophoresis buffer and heated to a high temperature until all of the agarose powder has melted.Smaller fragments of DNA are separated on higher concentrations of agarose whilst larger molecules require a lower concentration of agarose. The higher the agarose concentration, the denser the matrix and vice versa.The concentration of agarose used to make the gel depends on the size of the DNA fragments you are working with. Agarose gels are typically used to visualise fragments of DNA.How is gel electrophoresis carried out? Preparing the gel By comparing the bands of the DNA samples with those from the DNA marker, you can work out the approximate length of the DNA fragments in the samples.A DNA marker with fragments of known lengths is usually run through the gel at the same time as the samples.The use of dyes, fluorescent tags or radioactive labels enables the DNA on the gel to be seen after they have been separated. Shorter strands of DNA move more quickly through the gel than longer strands resulting in the fragments being arranged in order of size.DNA is negatively charged, therefore, when an electric current is applied to the gel, DNA will migrate towards the positively charged electrode.Electrophoresis enables you to distinguish DNA fragments of different lengths.As a result the molecules are separated by size. Smaller molecules migrate through the gel more quickly and therefore travel further than larger fragments that migrate more slowly and therefore will travel a shorter distance.The gel consists of a permeable matrix, a bit like a sieve, through which molecules can travel when an electric current is passed across it.A molecule with a negative charge will therefore be pulled towards the positive end (opposites attract!). Molecules migrate towards the opposite charge. The movement of charged molecules is called migration.An electric current is applied across the gel so that one end of the gel has a positive charge and the other end has a negative charge.Charged molecules move through a gel when an electric current is passed across it.Gel electrophoresis is a technique commonly used in laboratories to separate charged molecules like DNA, RNA and proteins according to their size.
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