Separation and isolation of DNA fragments in Recombinant DNA Technology


Gel electrophoresis
• After the cutting of DNA by restriction enzymes, fragments of DNA are formed. These fragments can be separated by a technique called gel electrophoresis.
• Electrophoresis is a technique of separation of charged molecules, under the influence of an electrical field, so that they migrate in the direction of electrode bearing the opposite charge, through a medium/matrix.
• The most commonly used matrix is agarose, which is a polysaccharide extracted from sea weeds.
• DNA fragments separate according to size, through the pores of agarose gel.
• The separated DNA fragments can be seen only after staining DNA with a compound ethidium bromide (EtBr), as bright orange coloured bands, on exposure to UV radiations.
• The separated bands of DNA are cut out from the agarose gel and extracted from the gel piece. This step is called as elution.

typical agarose gel electrophoresis

Cloning Vectors (Vehicle DNA)
• The vectors are DNA molecules that can carry a foreign DNA segment and replicate inside the host cell.

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