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 λbiot124-10 phage DNA and subsequently characterized by electron microscopic studies. The biotin-DNA fragment obtained after EcoRI cleavage of the λbiot124-10 DNA measures 18.7% λ DNA length (approx. 9000 base pairs). In addition to the biotin genes, it contains 4.75% and 3.08% λ phage DNA at the left and right endpoints 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 biotin-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.