Deoxyribonucleic Acid (DNA)

Restriction enzymes come from microorganisms. Recall that they recognize and cut DNA at specific base pair sequences. They cleave large DNA molecules into an assortment of smaller fragments ranging in size from a few to thousands of base pairs long, depending on how often and where the cleavage sequence appears in the original DNA molecule. The resulting fragments can be separated by their size using a technique known as electrophoresis. The fragments are placed at the top of a porous gel surrounded by a solution which conducts electricity. When a voltage is applied, the DNA fragments move towards the bottom of the gel due to the negative charge on their phosphate groups. Because it is more difficult for the large fragments to pass through the pores in the gel, they move more slowly than the smaller fragments.

DNA fragments isolated from a gel in this way can be joined with DNA from another source, either of the same or a different species, into a new, recombinant DNA molecule by enzymes. Usually, such DNA fragments are joined with DNA from subcellular organisms—"parasites" that live inside another organism but have their own DNA. Plasmids and viruses are two such examples. Viruses consist only of nucleic acids encapsulated in a protein coat. Though they can exist outside the cell, they are inactive. Inside the cell, they take over its metabolic machinery to manufacture more virus particles, eventually destroying their host. Plasmids are simpler than viruses in that they never exist outside the cell and have no protein coat. They consist only of circular double-stranded DNA. Plasmids replicate their DNA independently of their hosts. They are passed on to daughter cells in a controlled way as the host cell divides.

Cells that contain the same recombinant DNA fragment are clones. A clone harboring a recombinant DNA molecule that contains a specific gene can be isolated and identified by a number of techniques, depending upon the particular experiment. Thus, recombinant DNA molecules can be introduced into rapidly growing microorganisms, such as bacteria or yeast, to produce large quantities of medically or commercially important proteins normally present only in scant amounts in the cell. For example, human insulin and interferon have been produced in this manner.

In recent years a technique has been developed which permits analysis of very small samples of DNA without repeated cloning, which is laborious. Known as the polymerase chain reaction, this technique involves "amplifying" a particular fragment of DNA by repeated synthesis using the enzyme DNA polymerase. This method can increase the amount of the desired DNA fragment by a million-fold or more.