SUMMARY: Three sporulation mutants have been isolated which produce spores with an atypical resistance phenotype, i.e. they are sensitive to organic solvents and heat but resistant to lysozyme. All three mutants produced serine protease, alkaline phosphatase and glucose dehydrogenase which are biochemical marker events for stages I, II and III. Two of the three mutants produced dipicolinic acid, a late marker, but the third was defective in its production. Heat-resistance was not restored to any of the mutants by the provision of exogenous dipicolinate. Gel electrophoresis showed that the mutant spores had similar patterns of spore coat proteins to the wild-type and electron microscopy revealed no significant structural differences. The three mutations responsible for the phenotypes of the mutant spores lie in the phe-argA region of the Bacillus subtilis chromosome. Recombination index values indicate that the mutations are in three separate genes. They define at least two new sporulation loci, designated spoVH and spoVJ.
SUMMARY: λ;gt WES derivatives carrying the citG (fumarase) gene of Bacillus subtilis have been detected by complementation of an Escherichia coli fumarase mutation in lysogen-filled plaques. The gerA gene, mutations in which affect the germination response of spores to L-alanine, is also present on the two citG transducing phages described. The cloned regions in these two phages include EcoRI-generated fragments of 5.0 kb and 1.4 kb; the former fragment carries citG4 +.
SUMMARY: P1Kc-mediated transduction of Escherichia coli trp genes occurred at a frequency of about 10-8 in Salmonella typhimurium trp strains carrying mutations determining sensitivity to P1 and a low level of restriction enzymes. Heterospecific transductants were analysed by using them as donors in second-stage transductions mediated by bacteriophage KB int. One class of heterospecific transductants had the phenotype Trp+ Pro- but were extremely unstable and reverted at high frequency (up to 80%) to the parental phenotype. The Trp+ Pro- phenotype probably represents insertions of the E. coli trp genes in the S. typhimurium pro genes. It was stable in a RecA background.
SUMMARY: Mutants of Escherichia coli K12 defective in the gene iex (crr) no longer utilize glucose or N-acetylglucosamine in preference to lactose, but competition between either of these sugars and another that also enters by a phosphotransferase (PT) mechanism is not affected. In this they differ from gsr (tgs) mutants. In gsr mutants, glucose does not exclude any other sugar, though N-acetylglucosamine still does so. In gsr mutants that are also ptsM the phosphoenolpyruvate-dependent phosphorylation of glucose or methyl α-glucoside is reduced by 90%: N-acetylglucosamine phosphorylation is not affected. The iex mutation does not affect the phosphorylation of either of these compounds. The wild-type alleles iex + and gsr + are dominant in λ heterozygotes. Glucose inhibits the lactose permease of wild-type cells, but only when the permease is present in low amounts. The inhibition is also relieved (1) by induction of another transport system that is subject to regulation by the iex system or (2) by an iex mutation. We suggest that the iex gene specifies a protein that, in cells transporting certain sugars by a PT mechanism, acts to inhibit active transport systems. The protein is present in limiting concentration in the cell, sufficient only to inhibit the basal, uninduced, level of the active transport systems. In consequence the inducer (or its precursor) may be excluded from the cell and induction thus prevented.
SUMMARY: A lysogen of Escherichia coli K12 with λ;cI857 S7 xis6 nin5 b515 b519 integrated into ptsI was induced and the lysates plated on a Pel- host [on which λ; strains with less than the wild-type amount of DNA form plaques at low frequency (Cameron et al., 1977)]. All of the 40 plaques examined contained phage able to transduce at least two of the genes known from bacteriophage P1 transduction experiments to be closely linked to ptsI. Assuming that each specialized transducing phage arose by a single illegitimate recombination event, the distribution of phage types showed that the gene order is cysA gsr ptsI (ptsH, iex) cysZ lig; both gsr + and iex + were dominant. Analysis of restriction endonuclease digests of the transducing phage confirmed that no unexpected DNA rearrangements had taken place and allowed the construction of a map of the sites of action of the restriction endonucleases EcoRI, HindIII, BamI and Kpn for over 20 kilobases of E. coli DNA.
In an Appendix, we show cysA and cysZ mutants to be deficient in sulphate assimilation.
SUMMARY: Methods are described that allow extraction of high molecular weight DNA from germinated conidia of Neurospora crassa. By labelling DNA with ribonucleosides, early conidia were shown to be active in DNA synthesis. These cells when treated with the enzyme Zymolyase became fragile and could be readily lysed with ionic detergents to release high molecular weight DNA.
The DNA extracted from Zymolyase treated cells on to alkaline sucrose gradients sedimented as a heterogeneous species of up to 150×106 molecular weight. A minor DNA species (presumably mitochondrial) of 20×106 molecular weight comprised 2-7% of the total. The identity of the DNA was confirmed by sensitivity to DNAase, the diphenylamine assay and TLC. Sedimentation patterns were unaffected by protease digestions and no anomalous high speed rotor effects were evident. Isopycnic gradients suggested that the DNA released was uncomplexed with either protein or carbohydrates. Sepharose chromatography of extracted, RNAase-treated Zymolyase lysate resulted in clearly separate high molecular weight DNA and RNA-protein elution profiles.
UV light preferentially inhibited nuclear DNA synthesis and drastically reduced the size and amount of nascent DNA being synthesized in the excision defective uvs-2 mutant. Sites in parental DNA sensitive to Micrococous luteus UV endonuclease were measured in cells made permeable with Triton X-100.
SUMMARY: Streptomyces glaucescens strain GLA0 (= ETH 22794) produces hydroxystreptomycin and has a high natural resistance to hydroxystreptomycin, dihydrostreptomycin and streptomycin. The wild-type strain gives rise spontaneously to streptomycin-sensitive (StrS-) variants at a frequency of 0.2 to 1.4%. These mutants lack streptomycin phosphotransferase activity responsible for the wild-type resistance to streptomycin group antibiotics and are unable to produce detectable amounts of hydroxystreptomycin.
Mapping experiments showed that the strS marker lies between the chromosomal markers lys-2 and ura-3 on the linkage map of S. glaucescens. The molecular basis for instability of this marker is as yet unknown.