Search results for "DNA Primer"

Mutational Events in Cefotaximase Extended-Spectrum β-Lactamases of the CTX-M-1 Cluster Involved in Ceftazidime Resistance

2008

ABSTRACT CTX-M β-lactamases, which show a high cefotaxime hydrolytic activity, constitute the most prevalent extended-spectrum β-lactamase (ESBL) type found among clinical isolates. The recent explosive diversification of CTX-M enzymes seems to have taken place due to the appearance of more efficient enzymes which are capable of hydrolyzing both cefotaxime and ceftazidime, especially among the CTX-M-1 cluster. A combined strategy of in vitro stepwise evolution experiments using bla CTX-M-1 , bla CTX-M-3 , and bla CTX-M-10 genes and site-directed mutagenesis has been used to evaluate the role of ceftazidime and other β-lactam antibiotics in triggering the diversity found among enzymes belong…

Spatial and temporal changes in Actinobacterial dominance in experimental artificial groundwater recharge.

2008

Abstract Artificial groundwater recharge (AGR) is used in the drinking water industry to supplement groundwater resources and to minimise the use of chemicals in water treatment. This study analysed the spatial and temporal changes of microbial communities in AGR using two test systems: a nutrient-amended fluidized-bed reactor (FBR) and a sand column. Structural changes in the feed lake water (Lake Roine), FBR, and sand column bacterial communities were determined by denaturing gradient gel electrophoresis (DGGE) and the length heterogeneity analysis of amplified 16S rRNA genes (LH-PCR). Two clone libraries were created to link the LH-PCR results to the dominant bacterial groups. The lake w…

Cloning, deletion, and characterization of PadR, the transcriptional repressor of the phenolic acid decarboxylase-encoding padA gene of Lactobacillus…

2004

ABSTRACTLactobacillus plantarumdisplays a substrate-induciblepadAgene encoding a phenolic acid decarboxylase enzyme (PadA) that is considered a specific chemical stress response to the inducing substrate. The putative regulator ofpadAwas located in thepadAlocus based on its 52% identity with PadR, thepadAgene transcriptional regulator ofPediococcus pentosaceus(L. Barthelmebs, B. Lecomte, C. Diviès, and J.-F. Cavin, J. Bacteriol.182:6724-6731, 2000). Deletion of theL. plantarum padRgene clearly demonstrates that the protein it encodes is the transcriptional repressor of divergently orientedpadA. ThepadRgene is cotranscribed with a downstream open reading frame (ORF1), the product of which m…

A single primer pair gives a specific ortholog amplicon in a wide range of Cyanobacteria and plastid-bearing organisms: applicability in inventory of…

2010

The scarcity of universally applied molecular markers for algae has resulted in the development of multiple, independent and not easily comparable systems. The goal of this work is to increase the number of available molecular markers and to generate easily comparable systems. Thereby, we have designed a primer pair capable of amplifying a broad range of organisms: Cyanobacteria, Chlorophyta, Chlorarachniophyta, Cryptophyta, Euglenida, Glaucophyta, Rhodophyta, Stramenopiles and Streptophyta including plants. This primer pair can amplify a portion of the 23S rRNA gene with sufficient variability to identify reference material form collections across a broad range of taxa and perform phylogen…

PCR testing for Treponema pallidum in paraffin-embedded skin biopsy specimens: test design and impact on the diagnosis of syphilis

2007

Background: Syphilis, a chronic infection caused by Treponema pallidum (T. pallidum), is a disease which is increasing in incidence, and thus more and more becoming a differential diagnosis in routine pathology. Aim: Since histological changes are not specific, we sought to develop a polymerase chain reaction (PCR)-based molecular assay for the detection of T. pallidum in formalin-fixed, paraffin-embedded tissues, and evaluate its diagnostic power, especially in comparison with other ancillary methods, i.e. immunohistochemistry and Dieterle staining. Methods: 36 skin biopsies with the clinical and /or serological diagnosis of syphilis were evaluated by morphology, immunohistochemistry and s…

Cloning and characterization of the genes encoding the malolactic enzyme and the malate permease of Leuconostoc oenos

1996

Using degenerated primers from conserved regions of the protein sequences of malic enzymes, we amplified a 324-bp DNA fragment by PCR from Leuconostoc oenos and used this fragment as a probe for screening a Leuconostoc oenos genomic bank. Of the 2,990 clones in the genomic bank examined, 7 with overlapping fragments were isolated by performing colony hybridization experiments. Sequencing 3,453 bp from overlapping fragments revealed two open reading frames that were 1,623 and 942 nucleotides long and were followed by a putative terminator structure. The first deduced protein (molecular weight, 59,118) is very similar (level of similarity, 66%) to the malolactic enzyme of Lactococcus lactis; …

Nested PCR method for rapid and sensitive detection of Vibrio vulnificus in fish, sediments, and water

1995

A nested PCR for the detection of Vibrio vulnificus in fish farms was developed as an alternative to cultural methods by using universal primers flanking the V. vulnificus-specific sequences directed against 23S rRNA genes. This specific assay detected 10 fg of DNA or 12 to 120 cells in artificially inoculated samples without enrichment and within 24 h.

Rapid differentiation and in situ detection of 16 sourdough lactobacillus species by multiplex PCR.

2005

ABSTRACT A two-step multiplex PCR-based method was designed for the rapid detection of 16 species of lactobacilli known to be commonly present in sourdough. The first step of multiplex PCR was developed with a mixture of group-specific primers, while the second step included three multiplex PCR assays with a mixture of species-specific primers. Primers were derived from sequences that specify the 16S rRNA, the 16S-23S rRNA intergenic spacer region, and part of the 23S rRNA gene. The primer pairs designed were shown to exclusively amplify the targeted rrn operon fragment of the corresponding species. Due to the reliability of simultaneously identifying Lactobacillus plantarum , Lactobacillus…

Multiple Site-Specific Binding of Fis Protein to Escherichia coli nuoA-N Promoter DNA and its Impact on DNA Topology Visualised by Means of Scanning …

2004

Comparison of different primer sets for use in Automated Ribosomal Intergenic Spacer Analysis of complex bacterial communities.

2004

ABSTRACT ITSF and ITSReub, constituting a new primer set designed for the amplification of the 16S-23S rRNA intergenic transcribed spacers, have been compared with primer sets consisting of 1406F and 23Sr (M. M. Fisher and E. W. Triplett, Appl. Environ. Microbiol. 65:4630-4636, 1999) and S-D-Bact-1522-b-S-20 and L-D-Bact-132-a-A-18 (L. Ranjard et al., Appl. Environ. Microbiol. 67:4479-4487, 2001), previously proposed for automated ribosomal intergenic spacer analysis (ARISA) of complex bacterial communities. An agricultural soil and a polluted soil, maize silage, goat milk, a small marble sample from the façade of the Certosa of Pavia (Pavia, Italy), and brine from a deep hypersaline anoxi…