Restriction Enzymes Or Restriction Endonuclease

Genetic engineering is possible because of special enzymes that cut DNA. These enzymes are called restriction enzymes or restriction endonucleases. Restriction enzymes are proteins produced by bacteria to prevent or restrict invasion by foreign DNA. They act as DNA scissors, cutting the foreign DNA into pieces so that it cannot function. A nuclease is any enzyme that cuts the phosphodiester bonds of the DNA backbone, and an endonuclease is an enzyme that cuts somewhere within a DNA molecule. In contrast, an exonuclease cuts phosphodiester bonds by starting from a free end of the DNA and working inward. Restriction enzymes were originally discovered through their ability to break down, or restrict, foreign DNA. Restriction enzymes can distinguish between the DNA normally present in the cell and foreign DNA, such as infecting bacteriophage DNA. They defend the cell from invasion by cutting foreign DNA into

Also notice that some of the enzymes introduce two staggered cuts in the DNA, while others cut each strand at the same place. Enzymes like SmaI that cut both strands at the same place are said to produce blunt ends. Enzymes like EcoRI leave two identical DNA ends with single stranded protrusions:

5' G AATTC 3'

3' CTTAA G 5'

Under appropriate conditions (salt concentration, pH, and temperature), a given

Restriction enzyme will cleave a piece of DNA into a series of fragments. The number and sizes of the fragments depend on the number and location of restriction sites for that enzyme in the given DNA. A specific combination of bases will occur at random only once every few hundred bases, while a specific sequence of 6 will occur randomly only once every few thousand

Bases. It is possible that a DNA molecule will contain no restriction site for a given enzyme. For example, bacteriophage T7 (approximately 40,000 base pairs) contains no EcoRI