Isolation and characterization of the biotin genes of Escherichia coli K-12

Abstract

DNA containing the biotin gene cluster, bioABFCD, of E. coli K-12 has been isolated from the EcoRI cleavage products of lambdabiot124-10 phage DNA and subsequently characterized by electron microscopic studies. The biotin-DNA fragment obtained after EcoRI cleavage of the lambdabiot124-10 DNA measures 18.7% lambda DNA length (approx. 9000 base pairs). In addition to the biotin genes, it contains 4.75% and 3.08% lambda phage DNA at the left and right end-points of the bioABFCD cluster, respectively. The two bio promoter sites of the divergently transcribed biotin genes have been visualized under the electron microscope by binding RNA polymerase holoenzyme to the biotin DNA fragment. The two promoters are located at 41% and 43% length of the DNA fragment from its left endpoint. In vitro transcription of RNA from the bio-tin-DNA fragment has been visualized with the electron microscope, but so far no simultaneously transcribing "RNA:DNA" loops of the divergently oriented genes have been observed.