Gel Electrophoresis

Definition

A method used for the separation of deoxyribonucleic acid (DNA), ribonucleic acid (RNA), or protein molecules using an electric current applied to a gel matrix.

Application

Forensics, molecular biology experiment, genetics, microbiology and biochemistry

Difficulty

Procedure: Hard

Concept: Hard

This experiment contains toxic chemicals as well as extreme sterilization condition. Please refer to the Safety section for careful handling of chemicals.

Concept

Gel refers to a crosslinked polymer that can contain and separate a target molecule due to its pores inside. It is usually composed of different concentrations of acrylamide and a cross-linker, producing different mesh networks of polyacrylamide or agrose.

Electrophoresis refers to the electromotive force (EMF) that is required to move the molecules through the gel. Nucleic acids are negatively charged but placing them in a well that is on the negative side of the EMF will make them migrate towards the positive side of the gel.

Based on the size of the molecule, they will migrate at different rates. The lighter the molecule, the faster the molecule travels.

And in the case of nucleic acid, since it is one dimentional linear structure, the longer, or more base pairs one nucleic acid molecule contains, the heavier the molecule is. By letting different sample traveling over the same period of the time, they will stop at different locations. The distance travelled by the molecule is then inversely proportional to the mass of the molecule and can be used to determine what the unknown sample is by comparing its location to a unknown sample.

Materials

Procedure

Analysis

If automated softwares are available in conjunction with the UV camera, use the software to determine the size of the molecule by highlighting the band presented on the photo. The number of basepairs contained in each sample will be calculated and reported.

If automated softwares are not available, and accurate analysis is not required, the relative size of the molecule can be obtained by comparing the location of the band in the sample with the location of bands in the ladder. If the location is in the middle of two ladder bands, the number of basepairs in the sample band is then in the middle of those two ladder bands.

References