Separation and isolation of DNA fragments:
(i) The cutting of DNA by restriction endonucleases results in the short fragments of DNA, which can be separated by a technique known as gel electrophoresis.
(ii) The DNA fragments are negatively charged and they can be separated by forcing them to move towards the anode under an electric field through a medium/matrix.
(iii) Commonly used matrix is agarose, which is a natural linear polymer of D-galactose and 3, 6-anhydro-L-galactose which is extracted from sea weeds.
(iv) The DNA fragments separate (resolve) out according to their size through sieving effect provided by the agarose gel. Hence, the smaller the fragment size, the farther it will move.
(v) The separated DNA fragments can be visualised after staining the DNA with ethidium bromide followed by exposure to UV radiation.
(vi) The separated bands of DNA are cut out and extracted from the gel piece, this step is called elution.
(viii) The purified DNA fragments are used to form recombinant DNA which can be joined with cloning vectors.
