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The 54,383bp plasmid pKJK5 was recovered from a soil environment by exogenous plasmid isolation and conveys resistance towards tetracycline and trimethoprim. Sequencing and annotation revealed a high level of structural similarity of the backbone genes to other IncP-1 plasmids containing a Tra1 and Tra2 region, a central control module and a replication initiation module. A considerable degree of...
A novel Streptococcus pneumoniae plasmid (pSpnP1; 5413bp) has been isolated from the multidrug-resistant clone Poland 23F -16, and its complete nucleotide sequence has been determined. Sequence analysis predicted seven co-directional open reading frames and comparative analyses revealed that plasmid pSpnP1 is different to pDP1, the only previously described pneumococcal plasmid, whereas it...
Bacillus coagulans is a sporogenic lactic acid bacterium that ferments glucose and xylose, major components of plant biomass, a potential feedstock for cellulosic ethanol. The temperature and pH for optimum rate of growth of B. coagulans (50 to 55°C, pH 5.0) are very similar to that of commercially developed fungal cellulases (50°C; pH 4.8). Due to this match, simultaneous saccharification and fermentation...
A cryptic plasmid, pPG01 (3539bp), was isolated from Propionibacterium granulosum and sequenced. Analysis of open reading frames (ORFs) predicted pPG01 to encode three proteins. The largest protein (447 amino acids) showed homology to the FtsK/SpoIIIE family of proteins involved in chromosome partitioning during cell division and conjugal transfer of DNA. A second protein of 433 amino acids showed...
Previously reported techniques for the electrotransfer of foreign DNA into pediococci yield only a small number of transformants/μg DNA, especially when using undomesticated strains. This study reports an improved protocol for the electrotransformation of pediococci, based on trials using Pediococcus acidilactici P60 and the plasmid pRS4C1. The improved protocol yields from 2 to 3 log units more transformants...
The mobilizable transposon Tn4555, found in Bacteroides spp., is an important antibiotic resistance element encoding a broad spectrum β-lactamase. Tn4555 is mobilized by conjugative transposons such as CTn341 which can transfer the transposon to a wide range of bacterial species where it integrates into preferred sites on the host chromosome. Selection of the preferred target sites is mediated by...
Two novel plasmids, named pS4C and pL4C, were isolated from the thermophilic bacterium Thermus sp. 4C. The pS4C with a length of 5015bp and 58.25% of G+C content, contains 9 putative open reading frames (ORFs). The larger plasmid, pL4C, consisting of 21,248bp, has a G+C content of 68.60% and 34 putative ORFs. Both plasmids encode their own replication protein. The ORF 22 of pL4C and the ORF 4 of pS4C...
The antibiotic resistance plasmid pBS228 has been completely sequenced, and revealed to be descended from a plasmid virtually identical to the Birmingham IncP-1α plasmid RK2/RP4/RP1. However, it has three additional transposon insertions, one of which is responsible for the extra antibiotic resistances conferred. Loss of kanamycin resistance, which is characteristic of most IncP-1α plasmids, is the...
The genome of Epstein–Barr Virus (EBV) and plasmid derivatives of it are among the most efficient extrachromosomal replicons in mammalian cells. The latent origin of plasmid replication (oriP), when supplied with the viral Epstein–Barr Nuclear Antigen 1 (EBNA1) in trans, provides efficient duplication, partitioning and maintenance of plasmids bearing it. In this review, we detail what is known about...
In recent work, we described excision of a large genomic region from Enterococcus faecium D344R resulting from the interaction of Tn916 and a related transposon designated Tn5386. In the present study, we present and analyze the complete sequence of Tn5386. Tn5386 is 29,451 bp in length. Fifteen of its 30 open reading frames are analogous to ORFs found in Tn916. Significant differences include a series...
We report the characterization of two new theta-replicating plasmids of Streptococcus thermophilus (pSMQ-312b and pSMQ-316) as well as the further analysis of pSMQ-308. The nucleotide sequences of pSMQ-312b and pSMQ-316 were determined and both contained 6710 bp. In fact, the two sequences were identical, despite that the plasmids were isolated from two different S. thermophilus strains as demonstrated...
Recombineering, in vivo genetic engineering using the bacteriophage λ Red generalized recombination system, was used to create various modifications of a multicopy plasmid derived from pBR322. All genetic modifications possible on the Escherichia coli chromosome and on bacterial artificial chromosomes (BACs) are also possible on multicopy plasmids and are obtained with similar frequencies to their...
This work characterizes a recently discovered natural tetracycline-resistance plasmid called pMA67 from Paenibacillus larvae—a Gram-positive bacterial pathogen of honey bees. We provide evidence that pMA67 replicates by the rolling-circle mechanism, and sequence comparisons place it in the pMV158 family of rolling-circle replicons. The plasmid contains predicted rep, cop, and rnaII genes for control...
Francisella tularensis is a category A bioterror pathogen which in some cases can cause a severe and fatal human infection. Very few virulence factors are known in this species due to the difficulty in working with it as well as the lack of tools for genetic manipulation. This work describes the construction of a shuttle vector that can replicate in Escherichia coli and F. tularensis as well as two...
Plasmid pAM5 of Acidiphilium multivorum JCM-8867 has been completely sequenced by initial cloning of HindIII–PstI fragments followed by primer walking. It has a size of 5161bp and single site for several restriction enzymes as revealed by DNA sequencing. Sequence analysis predicts five putative open reading frames. ORF1 and ORF3 show significant identity with various plasmid encoded mobilization (Mob)...
Analysis by fluorescence microscopy has established that plasmid RK2 in Escherichia coli and other gram-negative bacteria is present as discrete clusters that are located inside the nucleoid at the mid- or quarter-cell positions. A mini-RK2 replicon containing an array of tetO repeats was visualized in E. coli cells that express a TetR-EYFP fusion protein. Unlike intact RK2, the RK2 mini-replicon...
Clostridium perfringens (C. perfringens) has been developed as a potential oral delivery vehicle to deliver antigens or therapeutic compounds to Gut-Associated Lymphoid Tissue (GALT). However, this recombinant C. perfringens carries a plasmid-encoded expression system, which raises several safety concerns regarding possible horizontal plasmid transfer and spread of plasmid-associated antibiotic resistant...
Bifidobacterium pseudolongum subsp. globosum DPC479 is an intestinally-derived strain which contains a plasmid, pASV479, 4.8kb in size. This plasmid has a G+C content of 59% and contains six open reading frames (ORFs), four of which are cryptic. The other two ORFs have 47% and 54% identity, respectively, to the replication and FtsK-like proteins found in a Bifidobacterium breve NCFB 2258 plasmid,...
Small interference RNA (siRNA) is a powerful tool for disrupting expression of specific genes in a variety of cells. We have developed a vector, piMARK, which mediates expression of both small hairpin RNA (shRNA) and the blasticidin resistance (Bsr) protein fused with enhanced green fluorescent protein (EGFP), enabling rapid selection and identification of knockdown cells. Using this vector, we targeted...
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