Insertion and Isolation of
Plasmid
SDS Gel Electrophoresis
Theory: The most commonly used method for determining the size and amount of protein chains is called sodium dodecyl sulfate (SDS) gel electrophoresis. This method can also find protein subunits in the preparation of a protein. The theory behind this method involves several steps. First a chemical is used to remove (break) all the disulfide bonds. This in turn causes disruption of all noncovalent bonds and interactions. Upon completion of this, you are left with two unfolded protein chains because they have denatured. The polypeptide chains are then placed in a polyacrilimide matrix. As the molecules travel through the matrix they will experience greater friction and travel less than smaller polypeptides. The most common form of polyacrilimide gel forms on one slab. There is a stacking gel and a running gel. The polypeptides are first loaded onto the stacking gel, where the separation of sizes begins.