In the year 1963, the two enzymes responsible for restricting the growth of bacteriophage in Escherichia coli were isolated. One of these added methyl groups to DNA, while the other cut DNA. The later was called restriction endonuclease.
The first restriction endonuclease–Hind II, whose functioning depended on a specific DNA nucleotide sequence was isolated and
characterised five years later. It was found that Hind II always cut DNA molecules at a particular point by recognising a specific sequence of six base pairs. This specific base sequence is known as the recognition sequence for Hind II. Besides Hind II, today we know more
than 900 restriction enzymes that have been isolated from over 230 strains of bacteria each of which recognise different recognition sequences.
The convention for naming these enzymes is the first letter of the name comes from the genus and the second two letters come from the species of the prokaryotic cell from which they were isolated, e.g., EcoRI comes from Escherichia coli RY 13. In EcoRI, the letter ‘R’ is derived from the name of strain. Roman numbers following the names indicate the order in which the enzymes were isolated from that strain of bacteria.
Restriction enzymes belong to a larger class of enzymes called nucleases. These are of two kinds; exonucleases and endonucleases.
Exonucleases remove nucleotides from the ends of the DNA whereas, endonucleases make cuts at specific positions within the DNA.
You may have realised that normally, unless one cuts the vector and the source DNA with the same restriction enzyme, the recombinant vector molecule cannot be created.

The cutting of DNA by restriction endonucleases results in the fragments of DNA. These fragments can be separated by a technique known as gel electrophoresis. Since DNA fragments are negatively charged molecules they can be separated by forcing them to move towards the anode under an electric field through a medium/matrix. Nowadays the most commonly used matrix is agarose which is a natural polymer extracted from sea weeds. The DNA fragments separate (resolve) according to their size through sieving effect provided by the agarose gel. Hence, the smaller the fragment size, the farther it moves. Look at the Figure 11.3 and guess at which end of the gel the sample was loaded.
The separated DNA fragments can be visualised only after staining the DNA with a compound known as ethidium bromide followed by exposure to UV radiation (you cannot see pure DNA fragments in the visible light and without staining). You can see bright orange coloured bands of DNA in a ethidium bromide stained gel exposed to UV light. The separated bands of DNA are cut out from the agarose gel and extracted from the gel piece. This step is known as elution. The DNA fragments purified in this way are used in
constructing recombinant DNA by joining them with cloning vectors.