Search results for "plasmid"
showing 10 items of 327 documents
Extraintestinal pathogenic Escherichia coli sequence type 131 H30-R and H30-Rx subclones in retail chicken meat, Italy
2016
Extraintestinal pathogenic Escherichia coli sequence type 131 (ST131), typically fluoroquinolone-resistant (FQ-R) and/or extended-spectrum β-lactamase (ESBL)-producing, has emerged globally. Among clinical isolates, ST131, primarily its H30-R and H30-Rx subclones, accounts for most antimicrobial-resistant E. coli and is the dominant E. coli strain worldwide. We assessed its prevalence and characteristics among raw chicken meat samples on sale in Palermo, Italy. A collection of 237 fluoroquinolone resistant and ESBL/AmpC producing E. coli isolates, which had been isolated from processed retail chicken meat in the period May 2013-April 2015, was analyzed. Established polymerase chain reaction…
Role of the metalloprotease Vvp and the virulence plasmid pR99 of Vibrio vulnificus serovar E in surface colonization and fish virulence.
2007
The virulence for eels of Vibrio vulnificus biotype 2 serovar E (VSE) is conferred by a plasmid that codifies ability to survive in eel serum and cause septicaemia. To find out whether the plasmid and the selected chromosomal gene vvp plays a role in the initial steps of infection, the VSE strain CECT4999, the cured strain CT218 and the Vvp-deficient mutant CT201 (obtained in this work by allelic exchange) were used in colonization and virulence experiments. The eel avirulent biotype 1 (BT1) strain YJ016, whose genome has been sequenced, was used for comparative purposes. The global results demonstrate that the plasmid does not play a significant role in surface colonization because (i) CEC…
Establishment of two quantitative nested qPCR assays targeting the human EPO transgene.
2016
International audience; For ethical and safety reasons it is critical to develop easily implemented assays with high sensitivity and specificity for gene doping surveillance. Two nested quantitative real-time PCR (qPCR) assays were developed that target the human EPO (hEPO) cDNA sequence in a circular form, representative of recombinant adeno-associated viral (rAAV) vector genomes found in vivo. Through an interlaboratory evaluation, the assays were validated and utilized in an in vitro blinded study. These assays are specific and extremely sensitive with a limit of detection (LOD) of 1 copy of circular plasmid DNA and a limit of quantification (LOQ) of 10 to 20 copies in the presence of 50…
Inducible metabolism of phenolic acids in Pediococcus pentosaceus is encoded by an autoregulated operon which involves a new class of negative transc…
2000
ABSTRACTPediococcus pentosaceusdisplays a substrate-inducible phenolic acid decarboxylase (PAD) activity onp-coumaric acid. Based on DNA sequence homologies between the three PADs previously cloned, a DNA probe of theLactobacillus plantarum pdcgene was used to screen aP. pentosaceusgenomic library in order to clone the corresponding gene of this bacteria. One clone detected with this probe displayed a low PAD activity. Subcloning of this plasmid insertion allowed us to determine the part of the insert which contains a 534-bp open reading frame (ORF) coding for a 178-amino-acid protein presenting 81.5% of identity withL. plantarumPDC enzyme. This ORF was identified as thepadAgene. A second O…
Homemade Site Directed Mutagenesis of Whole Plasmids
2009
Site directed mutagenesis of whole plasmids is a simple way to create slightly different variations of an original plasmid. With this method the cloned target gene can be altered by substitution, deletion or insertion of a few bases directly into a plasmid. It works by simply amplifying the whole plasmid, in a non PCR-based thermocycling reaction. During the reaction mutagenic primers, carrying the desired mutation, are integrated into the newly synthesized plasmid. In this video tutorial we demonstrate an easy and cost effective way to introduce base substitutions into a plasmid. The protocol works with standard reagents and is independent from commercial kits, which often are very expensi…
The Candida albicans UBI3 gene encoding a hybrid ubiquitin fusion protein involved in ribosome biogenesis is essential for growth.
2003
We have constructed a conditional null mutant Candida albicans strain for the UBI3 gene which encodes a ubiquitin fusion protein involved in ribosome biogenesis. A one-step gene disruption procedure, using the plasmid pCaDis, was designed to place the second copy of the UBI3 gene under the control of the tightly regulated MET3 promoter in a C. albicans heterozygous strain (UBI3/Deltaubi3::hisG), previously isolated in the first step of the ura-blaster protocol. Analysis of the conditional null mutant in repressing and inducing conditions indicates that UBI3 is an essential gene whose expression is required for growth of C. albicans.
Presence of a capsule in Vibrio vulnificus biotype 2 and its relationship to virulence for eels
1993
Strains of Vibrio vulnificus biotype 2, isolated from internal organs of diseased European eels as pure cultures of opaque cells, together with some reference strains from Japanese eels, were used in this study. Spontaneous translucent-phase variants were obtained from the corresponding parent strains and compared for a variety of phenotypic traits related to virulence for eels. The rate of colony dissociation from opaque to translucent cells was higher (around 10(-2)) than that observed for translucent to opaque cells (10(-3) to 10(-4)). Electron microscopy with ruthenium red revealed the presence of a capsule of variable thickness on opaque cells, whereas translucent-type colonies had no …
In vivo assembly of chromatin on pBR322 sequences cloned into yeast plasmids
1989
Abstract In order to study the in vivo assembly of chromatin on prokaryotic DNA templates, we have transformed yeast cells with plasmids pAJ50 and pRB58, which contain pBR322 sequences. In both cases nucleosomes are assembled in vivo on pBR322 DNA, although the nucleosomes are not homogeneous in size. To explore whether there is any preference for nucleosome assembly along pBR322 sequences, we have used an indirect end labeling method. The results indicate that most nucleosomes are placed at random on pBR322, although the probability for histone octamers to interact with some short regions is somewhat reduced. These regions coincide with sequences in which the frequency distribution of nucl…
A new glucose-repressible gene identified from the analysis of chromatin structure in deletion mutants of yeast SUC2 locus.
1991
We have previously shown that some changes occur in the chromatin structure of the 3' flank of the yeast SUC2 gene in going from a repressed to an active state. In an attempt to find out the causes of these changes, we have carried out experiments in which mutant copies of SUC2 locus lacking either 5' or 3' flanks have been analysed for their transcriptional activity and chromatin structure. These experiments allowed us to discard any relationship between SUC2 transcription and chromatin changes within its 3'flank. Sequencing of this flank and mRNA analysis, however, resulted in the location of a putative peroxisomal 3-oxoacyl-CoA thiolase gene (POT1), which is repressible by glucose. The d…
Genomic analysis of the emergence and evolution of multidrug resistance during a Klebsiella pneumoniae outbreak including carbapenem and colistin res…
2014
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