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We have developed a rapid method to investigate the influence of chromosome supercoiling on gene expression in Escherichia coli K12. This method exploits the ability of the gem3 mutant of the bacteriophage Mu, even in the prophagic state in immune cells, to induce relaxation of the host chromosome. The experiments can thus be performed under physiological conditions, and without the use of the drugs. In theory, this method can be applied to any bacterial gene. Here, we report the results obtained with four DNA replication and three cell division genes. 相似文献
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A new type of haem-deficient mutant was isolated in Escherichia coli K12 by neomycin selection. The mutant was deficient in uroporphyrinogen III cosynthase activity as indicated by the accumulation of uroporphyrin I and coproporphyrin. The mapping of the corresponding hemD gene by P1-mediated transduction showed that the new gene was located between ilv and cya, at min 83 on the chromosomal map of Escherichia coli K12. 相似文献
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Daniel Dignard Malcolm Whiteway Doris Germain Daniel Tessier David Y. Thomas 《Molecular & general genetics : MGG》1991,227(1):127-136
Summary A cDNA copy of the M2 dsRNA encoding the K2 killer toxin ofSaccharomyces cerevisiae was expressed in yeast using the yeastADH1 promoter. This construct produced K2-specific killing and immunity functions. Efficient K2-specific killing was dependent
on the action of the KEX2 endopeptidase and the KEX1 carboxypeptidase, while K2-specific immunity was independent of these
proteases. Comparison of the K2 toxin sequence with that of the K1 toxin sequence shows that although they share a common
processing pathway and are both encoded by cytoplasmic dsRNAs of similar basic structure, the two toxins are very different
at the primary sequence level. Site-specific mutagenesis of the cDNA gene establishes that one of the two potential KEX2 cleavage
sites is critical for toxin action but not for immunity. Immunity was reduced by an insertion of two amino acids in the hydrophobic
amino-terminal region which left toxin activity intact, indicating an independence of toxin action and immunity. 相似文献
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The nature of the transformation process in Escherichia coli K12 总被引:29,自引:0,他引:29
Summary The nature of the transformation process in E. coli was studied by investigating various factors which affect the efficiency of transformation. CaCl2 treatment of the recipient cells is absolutely necessary for transformation and the optimum concentration was found to be 30 mM. The efficiency of transformation is dependent upon temperature during incubation of the recipient cells with DNA. The efficiency is also affected by the molecular weight of donor DNA used. Sheared DNA with molecular weights ranging from 10 to 30x106 daltons was most efficient, increasing the number of transformants by a factor of 5 to 10 as compared to unsheared DNA. The intracellular status of recB-recC DNase (ATP-dependent DNase) is another important factor which determines the transformability of E. coli K12. This was shown by demonstrating that a recB
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recC
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sbcA
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recC
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sbcB
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strain. Therefore, it seems reasonable to conclude that the E. coli K12 strain is transformable if the ATP-dependent DNase is absent or diminished in function and a state of recombinational proficiency exists. 相似文献
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Properties of Escherichia coli K12 mutants which show conditional refractivity to colicin E2 总被引:1,自引:0,他引:1
I B Holland 《Journal of molecular biology》1968,31(2):267-275
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An investigation of the kinetics of photoreactivation (PR) in stationary phase cells of the UV-sensitive Escherichia coli mutant K12 AB2480 revealed an initial fast rate of PR which can be attributed to the photolysis of photoreactivating enzyme-substrate complexes already in existence at the start of the illumination period. The initial fast rate could be eliminated by administering a single high intensity flash at the start of continuous illumination. This experimental technique allowed the recognition of 2 additional rate processes during continuous illumination. The first order rate constant for the faster rate process (kf) leads to a value of 4–5 photoreactivating enzyme (PRE) molecules per cell for 16 h nutrient broth grown cells. A photoreactivating irradiance of approx. 60 erg mm?2 sec?1 saturates the reaction and under saturating conditions we obtain a value for the activation energy of the fast rate of 9 kcal mole?1. 相似文献
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Summary A pleiotropic mutant of Escherichia coli K
12 lacking reduced NAD: nitrate oxidoreductase, soluble formate dehydrogenase and membrane-bound formate:ferricytochrome b1 oxidoreductase is described. Levels of several other enzymes and cytochromes have been measured and found to differ little from those normally present in the wild type with the exceptions of cytochrome c522, reduced NAD:cytochrome c oxidoreductase and reduced NAD:nitrite oxidoreductase which are very high. Although the affected gene maps in a different position from that reported for chl A by other workers it seems likely that the two loci are identical. 相似文献
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A mutant of Escherichia coli which accumulates a large amount of coproporphyrin, presumably because of a block in heme biosynthesis, has been isolated after nitrosogunidine mutagenesis. On rich media, the mutant forms colonies which give bright orange fluorescence when illuminated with ultraviolet light. The mutant appears to be similar to a Salmonella typhimurium mutant, deficient in uroporphyrinogen III cosynthase, described by Sasarman and Desrochers ((1976) J. Bacteriol. 128, 717–721). A striking property of the mutant is that coproporphyrin is retained within the cells in rich media but is almost totally excreted out of cells in minimal glucose medium. 相似文献
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In Escherichia coli, dihydrofolate reductase is required for both the de novo synthesis of tetrahydrofolate and the recycling of dihydrofolate produced during the synthesis of thymidylate. The coding region of the dihydrofolate reductase gene, folA, was replaced with a kanamycin resistance determinant. Unlike earlier deletions, this mutation did not disrupt flanking genes. When the mutation was transferred into a wild-type strain and a thymidine-(thy) requiring strain, the resulting strains were viable but slow growing on rich medium. Both synthesized less folate than their parents, as judged by the incorporation of radioactive para-aminobenzoic acid. The derivative of the wild-type strain did not grow on any defined minimal media tested. In contrast, the derivative of the thy-requiring strain grew slowly on minimal medium with thy but exhibited auxotrophies on some combinations of supplements. These results suggest that when folates are limited, they can be distributed appropriately to folate-dependent biosynthetic reactions only under some conditions. 相似文献
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Transport of three synthesized endothiopeptides: AlaPsi[CSNH]Ala, AlaPsi[CSNH]Leu, and AlaPsi[CSNH]Phe, into Escherichia coli K12 mutant strains and enzymatic degradation studies were carried out. These compounds, well transported by permeases, but significantly more resistant to enzymatic cleavage than the corresponding natural peptides, seem to be useful as enzyme inhibitor carriers. 相似文献
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Escherichia coli K-12 gyrB gene product is involved in the lethal effect of the ligts2 mutant of bacteriophage Mu. 总被引:3,自引:2,他引:1
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P Ghelardini J C Liebart C Marchelli A M Pedrini L Paolozzi 《Journal of bacteriology》1984,157(2):665-668
Mu ligts2 mutants, defective for development and integration, show a high killing effect on the infected host. A number of survivors to Mu ligts2 infection were analyzed; they are characterized by nonpermissivity for both development and lysogenization of bacteriophage Mu. Bacteriophages D108 and P1 are also inhibited in these strains as is transposon Tn9. The corresponding mutation site was mapped at 82 min and identified with the Escherichia coli gyrB site. 相似文献
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Vasilieva S 《Mutation research》2001,496(1-2):89-95
PABA - Vitamin H1 of group B, has obtained increasing fundamental interest as a very potent natural antimutagen after a series of our publications since 1979. In the first set of our experiments, we studied PABA in the assays with the alkylating agent N-methyl-N-nitrosourea (MNU). Mutagenic efficiency of this agent was suppressed up to 10-fold when PABA was administered into Escherichia coli cells concurrently with the mutagen or prior to the mutagenic treatment. NMR spectrometric and UV-spectrophotometric measurements did not reveal an interaction between the direct acting MNU and PABA, typical for some N-nitroso compounds and phenolics. PABA suppressed the error-prone DNA repair pathway induced by UV-irradiation. PABA decreased MNU-induced phage lambda lysogenic induction more than two orders of magnitude. PABA inhibited the thermal shift up to 400-fold in phage lambda from the permissive to non-permissive temperature in E. coli mutant tif-1 and decreased about two-fold W-reactivation of UV-damaged phage lambda. Chloramphenicol treatment of the cells just after the mutagenic treatment prevented the occurrence of PABA specific activity. The results suggest that PABA affects the SOS DNA repair pathway and the mutagenic response of E. coli. PABA appears to be an effective bioantimutagen reducing mutagenesis by modulating the error-prone DNA repair (SOS) response. 相似文献
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