scholarly journals The recovery of tryptophan A auxotrophs at high frequency in a strain of Salmonella typhimurium

1967 ◽  
Vol 10 (2) ◽  
pp. 127-134 ◽  
Author(s):  
S. Riyasaty ◽  
G. W. P. Dawson
Keyword(s):  
Salmonella Typhimurium ◽  
Anthranilic Acid ◽  
High Frequency ◽  
Minimal Medium ◽  
Genetic Change ◽  
Wild Type ◽  
In The Wild ◽  
Slow Growing

1. Auxotrophs were sought in a slow-growing reversion of the tryA47 strain of S. typhimurium. This reversion differs from tryA47 by a genetic change that is inseparable from the 47 site and has been designated 47S. Out of thirty-nine auxotrophs that derived from independent mutations, twenty-two grew on minimal medium supplemented with anthranilic acid. Eight of these auxotrophs were examined and each was shown to have the 47S site unchanged and to carry a further mutation in the tryA gene. These further mutations were shown to be at different sites in different auxotrophs.2. Auxotrophs were sought in the wild-type LT-2 strain. None out of thirty-six were mutant in the tryA gene.3. The 47S site in tryA47S was replaced by 47+ transduced from wild-type LT-2. Auxotrophs were sought in this strain and only one out of nineteen was mutant in the tryA gene.4. Auxotrophs were sought in a wild-type reversion of tryA47. Out of twenty-six none were mutant in the tryA gene.5. The 47+ site in wild-type LT-2 was replaced by 47S transduced from tryA47S. Auxotrophs were sought in this strain and twenty-two out of fifty-five were mutant in the tryA gene.6. We conclude that tryA auxotrophs are only recovered at a high frequency when the 47S site is present in the tryA gene.7. In strains with the 47S site in the tryA gene the frequency of auxotrophs that will grow on minimal medium supplemented with indole but not on minimal medium supplemented with anthranilic acid is appreciably higher than in those strains without the 47S site in the tryA gene. These auxotrophs are mutant in genes that are in the same operon as tryA.

scholarly journals A genetic study of primary and secondary reversions of some tryptophanA auxotrophs of Salmonella typhimurium

1967 ◽  
Vol 9 (3) ◽  
pp. 269-282 ◽  
Author(s):  
S. Riyasaty ◽  
G. W. P. Dawson
Keyword(s):  
Salmonella Typhimurium ◽  
Genetic Change ◽  
Wild Type ◽  
Genetic Changes ◽  
Suppressor Mutations ◽  
Mutant Site ◽  
Back Mutation ◽  
Image Position ◽  
The Right ◽  
Slow Growing

1. The linkage order of four tryA mutants of S. typhimurium, and cysB-12, is:Attempts to plot the position of tryA-50 were unsuccessful.2. Some of the reversions of tryA-8, tryA-47, tryA-56 and tryA-50 were analysed genetically; tryA-52 does not revert. All four auxotrophs gave reversions that were phenotypically and genetically indistinguishable from that expected by back-mutation of the original mutant site.3. Both tryA-8 and tryA-50 produced reversions that grew as wild-type but were due to unlinked suppressor mutations. Some of these were super-suppressors in that they suppressed both tryA-8 and tryB-4; others suppressed many site mutants in the tryA gene but did not suppress tryB-4.4. All the slow-growing reversions of tryA-8, tryA-50 and tryA-56, and a minority of the semi-fast reversions of tryA-8, were due to unlinked suppressors.5. All the slow-growing reversions of tryA-47, the semi-fast reversions of tryA-56 and the majority of the semi-fast reversions of tryA-8 were due to genetic changes that were inseparable, in very extensive experiments, from their original mutant site.6. Slow-growing reversions of tryA-47 produced faster growing mutants. Some of these were due to mutation in unlinked modifying genes and in others the genetic change was within the tryA gene. Nine of the latter had the genetic change just to the left of the 47S site; in one the change was inseparable from the 47S site. None had this further change to the right of the 47S site. These further changes, in the absence of the 47S site, gave prototrophic phenotypes; they are inter-site suppressors.

scholarly journals Demonstration of different mutational sites controlling rhamnose fermentation in FIRN and non-FIRN rha−strains ofSalmonella typhimurium: an essay in bacterial archaeology

1968 ◽  
Vol 11 (2) ◽  
pp. 151-169 ◽  
Author(s):  
A. Morgenroth ◽  
J. P. Duguid
Keyword(s):  
Salmonella Typhimurium ◽  
Minimal Medium ◽  
Wild Type ◽  
Recombinant Bacteria ◽  
Peptone Water ◽  
Minimal Media ◽  
Different Strains ◽  
Ancestral Strain

Observations of mutation and phage-mediated transduction of rhamnose-fermentation properties were made on two groups of wild-type rhamnose-non-fermenting (rha−) strains ofSalmonella typhimuriumisolated in several different countries. The ‘FIRN’ group included twenty-three strains in the biotypes 15, 16, 17 and 18, and the ‘non-FIRN rha−’ group included fourteen strains in a new biotype, provisionally designated 19 X d. The two groups showed different patterns of mutational behaviour and the pattern was uniform for the different strains in each group. Non-FIRN rha−strains gave rise directly to colonies of rha+mutant bacteria in platings on rhamnose minimal media. FIRN strains were unable to do this, but produced rha+bacteria by two successive mutations, the first of which yielded weakly fermenting (rhaw) bacteria capable of fermenting rhamnose in peptone water, but not of utilizing it for growth on minimal medium.Rha+recombinant bacteria were produced in transductional crosses between any one FIRN strain and any one non-FIRN rha−strain, but not in any cross between one FIRN strain and another FIRN strain or between one non-FIRN rha−strain and another non-FIRN rha−strain. These findings suggest that although the sites of the rha−mutations are different between the FIRN and non-FIRN rha−groups, they are located at identical sites on the chromosome in all strains in either group. It is concluded that all FIRN strains have probably descended from a single ancestral strain which underwent mutations at two sites in the rhamnose region of the chromosome and that all non-FIRN rha−strains have descended from a single ancestral bacterium which underwent a mutation at a third site in the rhamnose region.

scholarly journals A Combination of Three Mutations, dcd,pyrH, and cdd, Establishes Thymidine (Deoxyuridine) Auxotrophy in thyA+ Strains of Salmonella typhimurium

1998 ◽  
Vol 180 (22) ◽  
pp. 5891-5895 ◽  
Author(s):  
Nevan J. Krogan ◽  
Michelle L. Zaharik ◽  
Jan Neuhard ◽  
Rod A. Kelln
Keyword(s):  
Salmonella Typhimurium ◽  
Genetic Background ◽  
Kinase Activity ◽  
Null Mutation ◽  
Wild Type ◽  
Gene Encoding ◽  
Deoxycytidine Deaminase ◽  
In The Wild ◽  
Ump Kinase ◽  
Thymidine Auxotrophy

ABSTRACT The dum gene of Salmonella typhimurium was originally identified as a gene involved in dUMP synthesis (C. F. Beck et al., J. Bacteriol. 129:305–316, 1977). In the genetic background used in their selection, the joint acquisition of adcd (dCTP deaminase) and a dum mutation established a condition of thymidine (deoxyuridine) auxotrophy. In this study, we show that dum is identical to pyrH, the gene encoding UMP kinase. The level of UMP kinase activity in thedum mutant was found to be only 30% of that observed for the dum + strain. Thymidine prototrophy was restored to the original dum dcd mutant (KP1361) either by transduction using a pyrH + donor or by complementation with either of twopyrH +-carrying plasmids. Thymidine auxotrophy could be reconstructed in the dum + derivative (KP1389) by the introduction of a mutant pyrH allele. To define the minimal mutational complement necessary to produce thymidine auxotrophy in thyA + strains, adcd::Km null mutation was constructed. In the wild-type background, dcd::Km alone or in combination with a pyrH (dum) mutation did not result in a thymidine requirement. A third mutation, cdd(cytidine-deoxycytidine deaminase), was required together with thedcd and pyrH mutations to impart thymidine auxotrophy.

scholarly journals Listeria monocytogenes PerR Mutants Display a Small-Colony Phenotype, Increased Sensitivity to Hydrogen Peroxide, and Significantly Reduced Murine Virulence

2005 ◽  
Vol 71 (12) ◽  
pp. 8314-8322 ◽  
Author(s):  
Rosemarie Rea ◽  
Colin Hill ◽  
Cormac G. M. Gahan
Keyword(s):  
Listeria Monocytogenes ◽  
High Frequency ◽  
Transcriptional Analysis ◽  
Wild Type ◽  
Large Colony ◽  
In The Wild ◽  
Increased Sensitivity ◽  
Small Colony

ABSTRACT Deletion of perR in Listeria monocytogenes results in a small-colony phenotype (ΔperR sm) that is slow growing and exhibits increased sensitivity to H2O2. At a relatively high frequency, large-colony variants (ΔperR lg) arise, which are more resistant to H2O2 than the wild-type and ultimately dominate the culture. Transcriptional analysis revealed that the kat gene (catalase) is up-regulated in both types of mutants and that the highest level is apparent in ΔperR sm mutants, demonstrating PerR regulation of this gene. Overexpression of the catalase gene in the wild-type background resulted in a slower-growing strain with a smaller colony size similar to that of ΔperR sm. By combining a bioinformatic approach with experimental evidence, other PerR-regulated genes were identified, including fur, lmo0641, fri, lmo1604, hemA, and trxB. The transcriptional profile of these genes in both mutant backgrounds was similar to that of catalase in that a higher level of expression was observed in ΔperR sm than in the wild type or ΔperR lg. Murine studies revealed that the virulence potential of the ΔperR sm mutant is substantially reduced compared to that of the wild-type and ΔperR lg strains. Collectively, the data demonstrate that the ΔperR sm mutant represents the true phenotype associated with the absence of PerR, which is linked to overexpression of regulated genes that negatively affect bacterial homeostasis both in vitro and in vivo. A subsequent secondary mutation occurred at a high frequency, which resulted in phenotypic reversion to a large-colony phenotype with increased fitness that may have obstructed the analysis of the role of PerR in the physiology of the bacterial cell.

scholarly journals Cloning of the chromosomal determinants encoding hemolysin production and mannose-resistant hemagglutination in Escherichia coli

1982 ◽  
Vol 152 (3) ◽  
pp. 1241-1247
Author(s):  
H Berger ◽  
J Hacker ◽  
A Juarez ◽  
C Hughes ◽  
W Goebel
Keyword(s):  
Escherichia Coli ◽  
High Frequency ◽  
Wild Type ◽  
Chromosomal Dna ◽  
Genetic Determinants ◽  
E Coli ◽  
Gene Banks ◽  
In The Wild ◽  
K 12 ◽  
Type Strains

We have cloned the chromosomal hemolysin determinants from Escherichia coli strains belonging to the four O-serotypes O4, O6, O18, and O75. The hemolysin-producing clones were isolated from gene banks of these strains which were constructed by inserting partial Sau3A fragments of chromosomal DNA into the cosmid pJC74. The hemolytic cosmid clones were relatively stable. The inserts were further subcloned either as SalI fragments in pACYC184 or as BamHI-SalI fragments in a recombinant plasmid (pANN202) containing cistron C (hlyC) of the plasmid-encoded hemolysin determinant. Detailed restriction maps of each of these determinants were constructed, and it was found that, despite sharing overall homology, the determinants exhibited minor specific differences in their structure. These appeared to be restricted to cistron A (hlyA), which is the structural gene for hemolysin. In the gene banks of two of these hemolytic strains, we could also identify clones which carried the genetic determinants for the mannose-resistant hemagglutination antigens Vb and VIc. Both of these fimbrial antigens were expressed in the E. coli K-12 clones to an extent similar to that observed in the wild-type strains. These recombinant cosmids were rather unstable, and, in the absence of selection, segregated at a high frequency.

scholarly journals Generation of Iron-Independent Siderophore-Producing Agaricus bisporus through the Constitutive Expression of hapX

Genes ◽  
2021 ◽  
Vol 12 (5) ◽  
pp. 724
Author(s):  
Min-Seek Kim ◽  
Hyeon-Su Ro
Keyword(s):  
Agaricus Bisporus ◽  
Recombinant Strain ◽  
Minimal Medium ◽  
Binary Vector ◽  
Constitutive Expression ◽  
Pcr Analysis ◽  
Mrna Levels ◽  
Wild Type ◽  
Chromosomal Dna ◽  
In The Wild

Agaricus bisporus secretes siderophore to uptake environmental iron. Siderophore secretion in A. bisporus was enabled only in the iron-free minimal medium due to iron repression of hapX, a transcriptional activator of siderophore biosynthetic genes. Aiming to produce siderophore using conventional iron-containing complex media, we constructed a recombinant strain of A. bisporus that escapes hapX gene repression. For this, the A. bisporushapX gene was inserted next to the glyceraldehyde 3-phosphate dehydrogenase promoter (pGPD) in a binary vector, pBGgHg, for the constitutive expression of hapX. Transformants of A. bisporus were generated using the binary vector through Agrobacterium tumefaciens-mediated transformation. PCR and Northern blot analyses of the chromosomal DNA of the transformants confirmed the successful integration of pGPD-hapX at different locations with different copy numbers. The stable integration of pGPD-hapX was supported by PCR analysis of chromosomal DNA obtained from the 20 passages of the transformant. The transformants constitutively over-expressed hapX by 3- to 5-fold and sidD, a key gene in the siderophore biosynthetic pathway, by 1.5- to 4-fold in mRNA levels compared to the wild-type strain (without Fe3+), regardless of the presence of iron. Lastly, HPLC analysis of the culture supernatants grown in minimal medium with or without Fe3+ ions presented a peak corresponding to iron-chelating siderophore at a retention time of 5.12 min. The siderophore concentrations of the transformant T2 in the culture supernatant were 9.3-fold (−Fe3+) and 8-fold (+Fe3+) higher than that of the wild-type A. bisporus grown without Fe3+ ions, while no siderophore was detected in the wild-type supernatant grown with Fe3+. The results described here demonstrate the iron-independent production of siderophore by a recombinant strain of A. bisporus, suggesting a new application for mushrooms through molecular biological manipulation.

Freeze-substituted fungal cells of Nectria haematococca: A comparison of the macroconidial walls of the adhesion-competent wild type with an adhesion-reduced mutant

1990 ◽  
Vol 48 (3) ◽  
pp. 688-689
Author(s):  
Thecan Caesar-Ton That ◽  
Lynn Epstein
Keyword(s):  
Freeze Substitution ◽  
Uranyl Acetate ◽  
Wild Type ◽  
Nectria Haematococca ◽  
Fruit Extract ◽  
Dialysis Tubing ◽  
Tube Emergence ◽  
Contrast Microscopy ◽  
In The Wild ◽  
Freeze Damage

Nectria haematococca mating population I (anamorph, Fusarium solani) macroconidia attach to its host (squash) and non-host surfaces prior to germ tube emergence. The macroconidia become adhesive after a brief period of protein synthesis. Recently, Hickman et al. (1989) isolated N. haematococca adhesion-reduced mutants. Using freeze substitution, we compared the development of the macroconidial wall in the wild type in comparison to one of the mutants, LEI.Macroconidia were harvested at 1C, washed by centrifugation, resuspended in a dilute zucchini fruit extract and incubated from 0 - 5 h. During the incubation period, wild type macroconidia attached to uncoated dialysis tubing. Mutant macroconidia did not attach and were collected on poly-L-lysine coated dialysis tubing just prior to freezing. Conidia on the tubing were frozen in liquid propane at 191 - 193C, substituted in acetone with 2% OsO4 and 0.05% uranyl acetate, washed with acetone, and flat-embedded in Epon-Araldite. Using phase contrast microscopy at 1000X, cells without freeze damage were selected, remounted, sectioned and post-stained sequentially with 1% Ba(MnO4)2 2% uranyl acetate and Reynold’s lead citrate. At least 30 cells/treatment were examined.

scholarly journals The Lipid Lysyl-Phosphatidylglycerol Is Present in Membranes of Rhizobium tropici CIAT899 and Confers Increased Resistance to Polymyxin B Under Acidic Growth Conditions

2007 ◽  
Vol 20 (11) ◽  
pp. 1421-1430 ◽  
Author(s):  
Christian Sohlenkamp ◽  
Kanaan A. Galindo-Lagunas ◽  
Ziqiang Guan ◽  
Pablo Vinuesa ◽  
Sally Robinson ◽  
...  
Keyword(s):  
Sinorhizobium Meliloti ◽  
Minimal Medium ◽  
Polymyxin B ◽  
Growth Conditions ◽  
Membrane Lipid ◽  
Low Ph ◽  
Wild Type ◽  
Rhizobium Tropici ◽  
Gram Negative ◽  
Inducible Genes

Lysyl-phosphatidylglycerol (LPG) is a well-known membrane lipid in several gram-positive bacteria but is almost unheard of in gram-negative bacteria. In Staphylococcus aureus, the gene product of mprF is responsible for LPG formation. Low pH-inducible genes, termed lpiA, have been identified in the gram-negative α-proteobacteria Rhizobium tropici and Sinorhizobium medicae in screens for acid-sensitive mutants and they encode homologs of MprF. An analysis of the sequenced bacterial genomes reveals that genes coding for homologs of MprF from S. aureus are present in several classes of organisms throughout the bacterial kingdom. In this study, we show that the expression of lpiA from R. tropici in the heterologous hosts Escherichia coli and Sinorhizobium meliloti causes formation of LPG. A wild-type strain of R. tropici forms LPG (about 1% of the total lipids) when the cells are grown in minimal medium at pH 4.5 but not when grown in minimal medium at neutral pH or in complex tryptone yeast (TY) medium at either pH. LPG biosynthesis does not occur when lpiA is deleted and is restored upon complementation of lpiA-deficient mutants with a functional copy of the lpiA gene. When grown in the low-pH medium, lpiA-deficient rhizobial mutants are over four times more susceptible to the cationic peptide polymyxin B than the wild type.

scholarly journals Rad51-Independent Interchromosomal Double-Strand Break Repair by Gene Conversion Requires Rad52 but Not Rad55, Rad57, or Dmc1

2007 ◽  
Vol 28 (3) ◽  
pp. 897-906 ◽  
Author(s):  
Thomas J. Pohl ◽  
Jac A. Nickoloff
Keyword(s):  
Gene Conversion ◽  
Strand Break ◽  
Double Strand Break ◽  
Single Strand ◽  
Homology Search ◽  
Wild Type ◽  
Dsb Repair ◽  
Single Strand Annealing ◽  
In The Wild ◽  
Break Induced Replication

ABSTRACT Homologous recombination (HR) is critical for DNA double-strand break (DSB) repair and genome stabilization. In yeast, HR is catalyzed by the Rad51 strand transferase and its “mediators,” including the Rad52 single-strand DNA-annealing protein, two Rad51 paralogs (Rad55 and Rad57), and Rad54. A Rad51 homolog, Dmc1, is important for meiotic HR. In wild-type cells, most DSB repair results in gene conversion, a conservative HR outcome. Because Rad51 plays a central role in the homology search and strand invasion steps, DSBs either are not repaired or are repaired by nonconservative single-strand annealing or break-induced replication mechanisms in rad51Δ mutants. Although DSB repair by gene conversion in the absence of Rad51 has been reported for ectopic HR events (e.g., inverted repeats or between plasmids), Rad51 has been thought to be essential for DSB repair by conservative interchromosomal (allelic) gene conversion. Here, we demonstrate that DSBs stimulate gene conversion between homologous chromosomes (allelic conversion) by >30-fold in a rad51Δ mutant. We show that Rad51-independent allelic conversion and break-induced replication occur independently of Rad55, Rad57, and Dmc1 but require Rad52. Unlike DSB-induced events, spontaneous allelic conversion was detected in both rad51Δ and rad52Δ mutants, but not in a rad51Δ rad52Δ double mutant. The frequencies of crossovers associated with DSB-induced gene conversion were similar in the wild type and the rad51Δ mutant, but discontinuous conversion tracts were fivefold more frequent and tract lengths were more widely distributed in the rad51Δ mutant, indicating that heteroduplex DNA has an altered structure, or is processed differently, in the absence of Rad51.

scholarly journals Effects of Mutations of RAD50, RAD51, RAD52, and Related Genes on Illegitimate Recombination in Saccharomyces cerevisiae

Genetics ◽  
1996 ◽  
Vol 142 (2) ◽  
pp. 383-391 ◽  
Author(s):  
Yasumasa Tsukamoto ◽  
Jun-ichi Kato ◽  
Hideo Ikeda
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Quantitative Analysis ◽  
Structural Analysis ◽  
Recombination Rate ◽  
Type Strain ◽  
Negative Selection ◽  
Wild Type Strain ◽  
Illegitimate Recombination ◽  
Wild Type ◽  
In The Wild

Abstract To examine the mechanism of illegitimate recombination in Saccharomyces cerevisiae, we have developed a plasmid system for quantitative analysis of deletion formation. A can1 cyh2 cell carrying two negative selection markers, the CAN1 and CYH2 genes, on a YCp plasmid is sensitive to canavanine and cycloheximide, but the cell becomes resistant to both drugs when the plasmid has a deletion over the CAN1 and CYH2 genes. Structural analysis of the recombinant plasmids obtained from the resistant cells showed that the plasmids had deletions at various sites of the CAN1-CYH2 region and there were only short regions of homology (1-5 bp) at the recombination junctions. The results indicated that the deletion detected in this system were formed by illegitimate recombination. Study on the effect of several rad mutations showed that the recombination rate was reduced by 30-, 10-, 10-, and 10-fold in the rad52, rad50, mre11, and xrs2 mutants, respectively, while in the rud51, 54, 55, and 57 mutants, the rate was comparable to that in the wild-type strain. The rad52 mutation did not affect length of homology at junction sites of illegitimate recombination.

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