scholarly journals Resistance ofEscherichia colito penicillins: II. An improved mapping of theampAgene

1968 ◽  
Vol 12 (2) ◽  
pp. 147-156 ◽  
Author(s):  
Kerstin G. Eriksson-Grennberg
Keyword(s):  
Escherichia Coli ◽  
Gene Order ◽  
Time Scale ◽  
Phenotypic Expression ◽  
Ampicillin Resistance ◽  
Ampicillin Concentration ◽  
Probable Gene

The first mutation step towards ampicillin resistance inEscherichia colioccurs in theampAgene, and gives resistance to a D, L-ampicillin concentration of 10 μg/ml. Using interrupted conjugation and transduction experimentsampAwas found to be located at 82 min on the time scale of Taylor & Thoman (1964). A number of adjacent markers were studied and the probable gene order of theampAregion was found to beargH-metA-uvrA-ampA-purA-fdp-pyrB. Two independent alleles ofampAwere cotransduced withfdp+andpurA+. The phenotypic expression ofampAin apurAstrain has been investigated.

Large-scale production of citrate synthase from a cloned gene

1982 ◽  
Vol 60 (12) ◽  
pp. 1143-1147 ◽  
Author(s):  
Harry W. Duckworth ◽  
Alexander W. Bell
Keyword(s):  
Escherichia Coli ◽  
Large Scale ◽  
Citrate Synthase ◽  
Specific Activity ◽  
Tetracycline Resistance ◽  
Scale Production ◽  
Ampicillin Resistance ◽  
Colicin E1 ◽  
Large Scale Production ◽  
Cloned Gene

Starting with a colicin E1 resistance recombinant plasmid which contains gltA, the gene for citrate synthase in Escherichia coli, we have constructed an ampicillin-resistance plasmid containing the gltA region as a 2.9-kilobase-pair insert in the tetracycline-resistance region of pBR322. Escherichia coli HB101 harbouring this plasmid, when grown on rich medium containing ampicillin, contains citrate synthase as about 8% of its soluble protein. The enzyme has been purified from this rich source and is identical to the chromosomal enzyme prepared previously in every property tested, except for specific activity, which is 64 U∙mg−1 as compared with 45–50 U∙mg−1 previously obtained. The N-terminal sequences of both enzymes are reported, and they are identical up to residue 16 at least. The overall yield of pure enzyme, starting with the cells grown in 15 L of medium, is 600–800 mg.

scholarly journals Stochasticity in Protein Levels Drives Colinearity of Gene Order in Metabolic Operons of Escherichia coli

PLoS Biology ◽  
2009 ◽  
Vol 7 (5) ◽  
pp. e1000115 ◽  
Author(s):  
Károly Kovács ◽  
Laurence D. Hurst ◽  
Balázs Papp
Keyword(s):  
Escherichia Coli ◽  
Gene Order ◽  
Protein Levels

scholarly journals A Combination of sbmA and tolC Mutations in Escherichia coli K-12 Tn10-Carrying Strains Results in Hypersusceptibility to Tetracycline

2007 ◽  
Vol 190 (4) ◽  
pp. 1491-1494 ◽  
Author(s):  
Ricardo E. de Cristóbal ◽  
Paula A. Vincent ◽  
Raúl A. Salomón
Keyword(s):  
Escherichia Coli ◽  
Double Mutant ◽  
Phenotypic Expression ◽  
Tetracycline Resistance ◽  
Additive Effect ◽  
Complete Suppression ◽  
High Level ◽  
K 12 ◽  
Level Resistance

ABSTRACT Previously, we demonstrated that Escherichia coli tolC mutations reduce the high-level resistance to tetracycline afforded by the transposon Tn10-encoded TetA pump from resistance at 200 μg/ml to resistance at 40 μg/ml. In this study, we found that the addition of an sbmA mutation to a tolC::Tn10 mutant exacerbates this phenotype: the double mutant did not form colonies, even in the presence of tetracycline at a concentration as low as 5 μg/ml. Inactivation of sbmA alone partially inhibited high-level tetracycline resistance, from resistance at 200 μg/ml to resistance at 120 μg/ml. There thus appears to be an additive effect of the mutations, resulting in almost complete suppression of the phenotypic expression of Tn10 tetracycline resistance.

scholarly journals Recovery of Plasmid pEMB1, Whose Toxin-Antitoxin System Stabilizes an Ampicillin Resistance-Conferring β-Lactamase Gene in Escherichia coli, from Natural Environments

2014 ◽  
Vol 81 (1) ◽  
pp. 40-47 ◽  
Author(s):  
Hyo Jung Lee ◽  
Hyun Mi Jin ◽  
Moon Su Park ◽  
Woojun Park ◽  
Eugene L. Madsen ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Plasmid Stability ◽  
Functional Characterization ◽  
Treatment Plant ◽  
Antibiotic Resistance Genes ◽  
Cloning And Expression ◽  
Natural Environments ◽  
Ampicillin Resistance ◽  
Content Type ◽  
Lactamase Gene

ABSTRACTNon-culture-based procedures were used to investigate plasmids showing ampicillin resistance properties in two different environments: remote mountain soil (Mt. Jeombong) and sludge (Tancheon wastewater treatment plant). Total DNA extracted from the environmental samples was directly transformed intoEscherichia coliTOP10, and a single and three different plasmids were obtained from the mountain soil and sludge samples, respectively. Interestingly, the restriction fragment length polymorphism pattern of the plasmid from the mountain soil sample, designated pEMB1, was identical to the pattern of one of the three plasmids from the sludge sample. Complete DNA sequencing of plasmid pEMB1 (8,744 bp) showed the presence of six open reading frames, including a β-lactamase gene. Using BLASTX, theorf5andorf6genes were suggested to encode a CopG family transcriptional regulator and a plasmid stabilization system, respectively. Functional characterization of these genes using a knockoutorf5plasmid (pEMB1ΔparD) and the cloning and expression oforf6(pET21bparE) indicated that these genes were antitoxin (parD) and toxin (parE) genes. Plasmid stability tests using pEMB1 and pEMB1ΔparDEinE. colirevealed that theorf5andorf6genes enhanced plasmid maintenance in the absence of ampicillin. Using a PCR-based survey, pEMB1-like plasmids were additionally detected in samples from other human-impacted sites (sludge samples) and two other remote mountain soil samples, suggesting that plasmids harboring a β-lactamase gene with a ParD-ParE toxin-antitoxin system occurs broadly in the environment. This study extends knowledge about the dissemination and persistence of antibiotic resistance genes in naturally occurring microbial populations.

scholarly journals Rearrangement of Gene Order in thephaCABOperon Leads to Effective Production of Ultrahigh-Molecular-Weight Poly[(R)-3-Hydroxybutyrate] in Genetically Engineered Escherichia coli

2012 ◽  
Vol 78 (9) ◽  
pp. 3177-3184 ◽  
Author(s):  
Ayaka Hiroe ◽  
Kenji Tsuge ◽  
Christopher T. Nomura ◽  
Mitsuhiro Itaya ◽  
Takeharu Tsuge
Keyword(s):  
Escherichia Coli ◽  
Molecular Weight ◽  
Gene Order ◽  
Ralstonia Eutropha ◽  
Genetically Engineered ◽  
Pha Synthase ◽  
Limiting Factor ◽  
Expression Plasmid ◽  
Content Type ◽  
Ultrahigh Molecular Weight

ABSTRACTUltrahigh-molecular-weight poly[(R)-3-hydroxybutyrate] [UHMW-P(3HB)] synthesized by genetically engineeredEscherichia coliis an environmentally friendly bioplastic material which can be processed into strong films or fibers. An operon of three genes (organized asphaCAB) encodes the essential proteins for the production of P(3HB) in the native producer,Ralstonia eutropha. The three genes of thephaCABoperon arephaC, which encodes the polyhydroxyalkanoate (PHA) synthase,phaA, which encodes a 3-ketothiolase, andphaB, which encodes an acetoacetyl coenzyme A (acetoacetyl-CoA) reductase. In this study, the effect of gene order of thephaCABoperon (phaABC,phaACB,phaBAC,phaBCA,phaCAB, andphaCBA) on an expression plasmid in genetically engineeredE. coliwas examined in order to determine the best organization to produce UHMW-P(3HB). The results showed that P(3HB) molecular weights and accumulation levels were both dependent on the order of thephagenes relative to the promoter. The most balanced production result was achieved in the strain harboring thephaBCAexpression plasmid. In addition, analysis of expression levels and activity for P(3HB) biosynthesis enzymes and of P(3HB) molecular weight revealed that the concentration of active PHA synthase had a negative correlation with P(3HB) molecular weight and a positive correlation with cellular P(3HB) content. This result suggests that the level of P(3HB) synthase activity is a limiting factor for producing UHMW-P(3HB) and has a significant impact on P(3HB) production.

scholarly journals Phenotypic expression in Escherichia coli and nucleotide sequence of two Chinese hamster lung cell cDNAs encoding different dihydrofolate reductases.

1984 ◽  
Vol 4 (1) ◽  
pp. 38-48 ◽  
Author(s):  
P W Melera ◽  
J P Davide ◽  
C A Hession ◽  
K W Scotto
Keyword(s):  
Escherichia Coli ◽  
Molecular Weight ◽  
Nucleotide Sequence ◽  
Dihydrofolate Reductase ◽  
Phenotypic Expression ◽  
Chinese Hamster ◽  
Differential Sensitivity ◽  
Nucleotide Sequence Analysis ◽  
Cdna Clones ◽  
Weight Class

Nucleotide sequence analysis of two cDNA clones, one shown to direct the synthesis in Escherichia coli of the pI 6.7 form of the 20,000-molecular-weight class of Chinese hamster lung cell dihydrofolate reductase, and the other shown to direct the synthesis of the pI 6.5 form of the 21,000-molecular-weight class of the enzyme, has revealed the following: (i) the differences in physical and enzymatic properties displayed by these two proteins are due to two variations in their respective amino acid sequences with the conversion of Leu to Phe at position 22 probably responsible for the differential sensitivity of these two enzymes to methotrexate and methasquin; (ii) the multiple mRNAs responsible for the synthesis of each of these proteins differ in size due, at least in part, to a length heterogeneity at their 3' ends; (iii) these two proteins are encoded by different genes; and (iv) the sequence AAATATA appears to be a major polyadenylation signal in one Chinese hamster lung cell dihydrofolate reductase gene and a minor signal in another.

scholarly journals Chromosomal location of a mutation causing chloramphenicol resistance in Escherichia coli K 12

1968 ◽  
Vol 11 (1) ◽  
pp. 97-104 ◽  
Author(s):  
E. C. R. Reeve ◽  
D. R. Suttie
Keyword(s):  
Escherichia Coli ◽  
Gene Order ◽  
Chromosomal Location ◽  
Attachment Site ◽  
Chloramphenicol Resistance ◽  
K 12 ◽  
Resistant Mutation

A chloramphenicol-resistant mutation in Escherichia coli K 12, cmlA1 (previously designated 1a), giving a higher Cm-resistance than other mutations yet examined, has been shown to have a chromosomal location, the gene order being gal, λ, bio, cmlA, pyrD. CmlA can be transduced efficiently into cm-sensitive strains by P1 with little phenotypic lag, and is co-transduced with the λ-attachment site (frequency 1·13%) but not with gal or pyrD.

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