Restriction enzyme cloning

Restriction enzyme based molecular cloning is the "classic" cloning method and is still one of the most popular techniques. In principle, the plasmid assembly involves these following steps:

• Restriction digest: Closed circular vector plasmid and gene of interest fragment are cleaved with one or more restriction enzymes.

• Ligation: DNA fragments with compatible overhangs are ligated into a linearized vector plasmid.

Higher success rates are achieved when using DNA fragments with 5' or 3' sticky/overhang ends rather than assembling DNA fragments with blunt ends.

• 3A assembly is a method used to efficiently assemble two BioBricks.