ESCHERICHIA-COLI RNA-POLYMERASE BINDING AND INITIATION OF TRANSCRIPTION ON FRAGMENTS OF LAMBDA-RIFD 18 DNA CONTAINING PROMOTERS FOR LAMBDA-GENES AND FOR RRNB, TUFB, RPLK,A, RPLJ,L, AND RPOB,C GENES

Promoters of genes for bacteriophage Λ and for Escherichia coli ribosomal RNA (rrnB), elongation factor Tu (tufB), ribosomal proteins L11 (rplK), L1 (rplA), L10 (rplJ), and L7/L12 (rplL), and RNA polymerase subunits β (rpoB) and β′ (rpoC) were studied by use of two types of filter binding assays which measured E. coli RNA polymerase binding and initiation of transcription on restriction fragments of Λrifd18 DNA. The DNA fragments selectively retained on filters were eluted, concentrated, and analyzed by gel electrophoresis. The binding characteristics of these promoter fragments were qualitatively determined by varying the RNA polymerase, salt, and glycerol concentrations in the polymerase binding assay with HaeIII fragments of Λrifd18 DNA. The approximate map locations of these small HaeIII fragments were determined by HaeIII digestion of the larger, previously mapped EcoRI, HindII, and SmaI restriction fragments of the phage DNA. The base compositions proximal to the 5′ ends of mRNA's from promoters on these DNA fragments were elucidated by the polymerase initiation assay, in which the addition of various combinations of nucleoside triphosphates to the reaction allowed RNA polymerase to form high-salt-resistant initiation complexes with some of the known SmaI + EcoRI, EcoRI + HindIII, or HaeIII restriction fragments of Λrifd18 DNA. The data obtained by this technique are consistent. © 1979.