Isolation and characterization of a ColE1 plasmid containing the entire Bio gene cluster of Escherichia coli K12

Abstract
ColE1 amp plasmids carrying the entire bio gene cluster were constructed in vitro using ColE1 amp as the cloning vehicle and a λ transducing phage, λatt2,as the source of bio DNA. Restriction endonuclease EcoRI digests of ColE1amp and λatt2DNA were joined by polynucleotide ligase and plasmids bearing the entire bio gene cluster were selected, after transformation, in bio deletion strains of E. coli. Recombinant DNA moleules contained one ColE1amp fragment (7.4×106 daltons) and one λatt2DNA fragment (5.4×106 daltons). Clones carrying ColE1 amp-bio plasmids produce elevated levels of biotin and biotin synthetase activity.