Once all the samples have been loaded into the wells, the chamber is connected to a power supply and an electrical current is applied to the gel. The chamber is designed with a positive electrode (anode) at one end and a negative electrode (cathode) at the other end. Molecules with a net negative charge migrate toward the positive electrode and molecules with a net positive charge migrate toward the negative electrode because opposite charges attract.
The overall charge of a molecule affects the speed at which it travels through the gel. Highly charged molecules migrate more quickly through the gel than weakly charged molecules. The size and shape of the molecule also affects how quickly it travels through the gel. Agarose gels contain a matrix of minuscule pores that acts like a sieve. Small molecules maneuver more easily through the pores of the gel than larger molecules, allowing them to migrate relatively quickly. Size and net charge are factors that together determine how quickly molecules will travel through the gel, and thus what their migration distance will be. If a molecule is small or highly charged, this will increase its migration rate through the gel. If a molecule is large or weakly charged, this will decrease its migration rate through the gel.
In this activity, five known dye samples and three unknown dye mixtures will be subjected to agarose gel electrophoresis. Some of the dyes will be attracted to the negative electrode and some to the positive electrode depending on their overall charge. Each of the known dyes will exhibit a unique gel migration distance that relates to its molecular size and net charge. You will identify the components of the unknown dye mixtures by comparing the migration distances and direction of migration of the unknown dyes to those of the known dye samples
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