Restriction enzymes (nucleases) are a class of enzymatic proteins that cut nucleotide chains at given locations or specific sequences. Restriction endonucleases are the most common form of restriction enzyme used in molecular cloning. Endonucleases recognize an internal sequence within a nucleotide chain, and cleave at that specific sequence generating either sticky ends (containing a 3’ or 5’ overhang) or blunt (containing no 3’ or 5’ overhang) ends. These ends may then be joined to a compatible end on another sequence via a DNA-ligase to prepare a desired sequence.
Restriction exonucleases do not recognize or cleave internal sequences on DNA, and instead function by removing nucleotides from a “free” or open end. Different exonucleases may work, either in a 5’ -> 3’ direction, or in a 3 -> 5’ direction. In molecular cloning, exonucleases are most commonly used to remove either a 3’ or 5’ overhang from a “sticky end” and leave, instead a “blunt” end (e.g. after a sequence has been cleaved with an endonuclease).
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