Browse
Submit Data
Databases
API
Help

Dataset Information

23 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Modification of RSF1010-Based Broad-Host-Range Plasmids for Improved Conjugation and Cyanobacterial Bioprospecting.

ABSTRACT: To facilitate the genetic engineering of diverse cyanobacterial strains, we have modified broad-host-range RSF1010-based plasmids to improve transmissibility, increase copy number, and facilitate cloning. RSF1010-based plasmids replicate in diverse bacterial strains but produce low amounts of useable DNA for cloning. We previously engineered a mobAY25F mutation in RSF1010-based plasmids that improved cloning but decreased conjugation efficiency. Here, we engineered RSF1010-based plasmids to restore conjugation efficiency, which was demonstrated in three diverse laboratory strains of cyanobacteria. We then used an improved RSF1010-based plasmid in mating experiments with cultured samples of wild cyanobacteria. This plasmid, which confers antibiotic resistance and carries a yfp reporter gene, allowed selection of exconjugant cyanobacteria and facilitated the isolation of genetically tractable strains from mixed wild cultures. Improved RSF1010 vectors can be used for bioprospecting genetically tractable strains and are compatible with the CYANO-VECTOR cloning system, a versatile toolbox for constructing plasmids for cyanobacterial genetic engineering.

SUBMITTER: Bishe B

PROVIDER: S-EPMC6817606 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

A relative of the broad-host-range plasmid RSF1010 detected in Erwinia amylovora.

Project description:Streptomycin- and sulfonamide-resistant Erwinia amylovora CA3R from California contained an 8.7-kb plasmid, pEa8.7, with a sulII-strA-strB resistance region; furthermore, PCR, sequencing, hybridization, and restriction analyses showed that pEa8.7 was closely related or identical to broad-host-range plasmid RSF1010. Although RSF1010 has been found in a variety of bacteria, this is the first report of its presence in plant pathogenic bacteria.

| S-EPMC168779 | biostudies-other

Broad-host-range IncP-1 plasmids and their resistance potential.

Project description:The plasmids of the incompatibility (Inc) group IncP-1, also called IncP, as extrachromosomal genetic elements can transfer and replicate virtually in all Gram-negative bacteria. They are composed of backbone genes that encode a variety of essential functions and accessory genes that have implications for human health and environmental bioremediation. Broad-host-range IncP plasmids are known to spread genes between distinct phylogenetic groups of bacteria. These genes often code for resistances to a broad spectrum of antibiotics, heavy metals, and quaternary ammonium compounds used as disinfectants. The backbone of these plasmids carries modules that enable them to effectively replicate, move to a new host via conjugative transfer and to be stably maintained in bacterial cells. The adaptive, resistance, and virulence genes are mainly located on mobile genetic elements integrated between the functional plasmid backbone modules. Environmental studies have demonstrated the wide distribution of IncP-like replicons in manure, soils and wastewater treatment plants. They also are present in strains of pathogenic or opportunistic bacteria, which can be a cause for concern, because they may encode multiresistance. Their broad distribution suggests that IncP plasmids play a crucial role in bacterial adaptation by utilizing horizontal gene transfer. This review summarizes the variety of genetic information and physiological functions carried by IncP plasmids, which can contribute to the spread of antibiotic and heavy metal resistance while also mediating the process of bioremediation of pollutants. Due to the location of the resistance genes on plasmids with a broad-host-range and the presence of transposons carrying these genes it seems that the spread of these genes would be possible and quite hazardous in infection control. Future studies are required to determine the level of risk of the spread of resistance genes located on these plasmids.

| S-EPMC3590792 | biostudies-literature

Diverse broad-host-range plasmids from freshwater carry few accessory genes.

Project description:Broad-host-range self-transferable plasmids are known to facilitate bacterial adaptation by spreading genes between phylogenetically distinct hosts. These plasmids typically have a conserved backbone region and a variable accessory region that encodes host-beneficial traits. We do not know, however, how well plasmids that do not encode accessory functions can survive in nature. The goal of this study was to characterize the backbone and accessory gene content of plasmids that were captured from freshwater sources without selecting for a particular phenotype or cultivating their host. To do this, triparental matings were used such that the only required phenotype was the plasmid's ability to mobilize a nonconjugative plasmid. Based on complete genome sequences of 10 plasmids, only 5 carried identifiable accessory gene regions, and none carried antibiotic resistance genes. The plasmids belong to four known incompatibility groups (IncN, IncP-1, IncU, and IncW) and two potentially new groups. Eight of the plasmids were shown to have a broad host range, being able to transfer into alpha-, beta-, and gammaproteobacteria. Because of the absence of antibiotic resistance genes, we resampled one of the sites and compared the proportion of captured plasmids that conferred antibiotic resistance to their hosts with the proportion of such plasmids captured from the effluent of a local wastewater treatment plant. Few of the captured plasmids from either site encoded antibiotic resistance. A high diversity of plasmids that encode no or unknown accessory functions is thus readily found in freshwater habitats. The question remains how the plasmids persist in these microbial communities.

| S-EPMC3837812 | biostudies-literature

Structure and function of primase RepB' encoded by broad-host-range plasmid RSF1010 that replicates exclusively in leading-strand mode.

Project description:For the initiation of DNA replication, dsDNA is unwound by helicases. Primases then recognize specific sequences on the template DNA strands and synthesize complementary oligonucleotide primers that are elongated by DNA polymerases in leading- and lagging-strand mode. The bacterial plasmid RSF1010 provides a model for the initiation of DNA replication, because it encodes the smallest known primase RepB' (35.9 kDa), features only 1 single-stranded primase initiation site on each strand (ssiA and ssiB, each 40 nt long with 5'- and 3'-terminal 6 and 13 single-stranded nucleotides, respectively, and nucleotides 7-27 forming a hairpin), and is replicated exclusively in leading strand mode. We present the crystal structure of full-length dumbbell-shaped RepB' consisting of an N-terminal catalytic domain separated by a long alpha-helix and tether from the C-terminal helix-bundle domain and the structure of the catalytic domain in a specific complex with the 6 5'-terminal single-stranded nucleotides and the C7-G27 base pair of ssiA, its single-stranded 3'-terminus being deleted. The catalytic domains of RepB' and the archaeal/eukaryotic family of Pri-type primases share a common fold with conserved catalytic amino acids, but RepB' lacks the zinc-binding motif typical of the Pri-type primases. According to complementation studies the catalytic domain shows primase activity only in the presence of the helix-bundle domain. Primases that are highly homologous to RepB' are encoded by broad-host-range IncQ and IncQ-like plasmids that share primase initiation sites ssiA and ssiB and high sequence identity with RSF1010.

| S-EPMC2683140 | biostudies-literature

The interaction of RepC initiator with iterons in the replication of the broad host-range plasmid RSF1010.

Project description:The replication origin of the broad host-range plasmid RSF1010 contains 3.5 copies of a 20mer iteron sequence that bind specifically to the plasmid-encoded initiator, RepC. Here we demonstrated that even a single iteron was bent upon binding of RepC. Moreover, the bending angle seems to become larger along with the increment of the number of iterons. In a mutational analysis of the iteron sequence, we isolated seven kinds of base-substitution mutants of iterons, and estimated the replication activity of these mutants in vivo. We found that each of the subsections in the 20mer iteron sequence made a distinct contribution to the initiation of RSF1010 DNA replication. With the binding assay of RepC and mutated iterons in vitro, we found that the formation of a productive RepC-iteron complex was required for the initiation of plasmid DNA replication.

| S-EPMC307191 | biostudies-other

Diversification of broad host range plasmids correlates with the presence of antibiotic resistance genes.

Project description:The IncP-1ε subgroup is a recently identified phylogenetic clade within IncP-1 plasmids, which plays an important role in the spread of antibiotic resistance and degradation of xenobiotic pollutants. Here, four IncP-1ε plasmids were exogenously captured from a petroleum-contaminated habitat in China and compared phylogenetically and genomically with previously reported IncP-1ε and other IncP-1 plasmids. The IncP-1ε plasmids can be clearly subdivided into two subclades, designated as ε-I and ε-II, based on phylogenetic analysis of backbone proteins TraI and TrfA. This was further supported by comparison of concatenated backbone genes. Moreover, the two subclades differed in the transposon types, phenotypes and insertion locations of the accessory elements. The accessory genes on ε-I plasmids were inserted between parA and traC, and harbored ISPa17 and Tn402-like transposon modules, typically carrying antibiotic resistance genes. In contrast, the accessory elements on ε-II plasmids were typically located between trfA and oriV, and contained IS1071, which was commonly inserted within the Tn501-like transposon, typically harboring a cluster of genes encoding mercury resistance and/or catabolic pathways. Our study is one of the first to compare IncP-1 plasmid genomes from China, expands the available collection of IncP-1ε plasmids and enhances our understanding of their diversity, biogeography and evolutionary history.

| S-EPMC4809973 | biostudies-literature

Genetic Characterization of Broad-Host-Range IncQ Plasmids Harboring blaVEB-18 in Vibrio Species.

Project description: Not available

| S-EPMC5487652 | biostudies-literature

Broad-host-range plasmids from agricultural soils have IncP-1 backbones with diverse accessory genes.

Project description:Broad-host-range plasmids are known to spread genes between distinct phylogenetic groups of bacteria. These genes often code for resistances to antibiotics and heavy metals or degradation of pollutants. Although some broad-host-range plasmids have been extensively studied, their evolutionary history and genetic diversity remain largely unknown. The goal of this study was to analyze and compare the genomes of 12 broad-host-range plasmids that were previously isolated from Norwegian soils by exogenous plasmid isolation and that encode mercury resistance. Complete nucleotide sequencing followed by phylogenetic analyses based on the relaxase gene traI showed that all the plasmids belong to one of two subgroups (? and ?) of the well-studied incompatibility group IncP-1. A diverse array of accessory genes was found to be involved in resistance to antimicrobials (streptomycin, spectinomycin, and sulfonamides), degradation of herbicides (2,4-dichlorophenoxyacetic acid and 2,4-dichlorophenoxypropionic acid), and a putative new catabolic pathway. Intramolecular transposition of insertion sequences followed by deletion was found to contribute to the diversity of some of these plasmids. The previous observation that the insertion sites of a Tn501-related element are identical in four IncP-1? plasmids (pJP4, pB10, R906, and R772) was further extended to three more IncP-1? plasmids (pAKD15, pAKD18, and pAKD29). We proposed a hypothesis for the evolution of these Tn501-bearing IncP-1? plasmids that predicts recent diversification followed by worldwide spread. Our study increases the available collection of complete IncP-1 plasmid genome sequences by 50% and will aid future studies to enhance our understanding of the evolution and function of this important plasmid family.

| S-EPMC3209000 | biostudies-literature

Plasmids do not consistently stabilize cooperation across bacteria but may promote broad pathogen host-range.

Project description:Horizontal gene transfer via plasmids could favour cooperation in bacteria, because transfer of a cooperative gene turns non-cooperative cheats into cooperators. This hypothesis has received support from theoretical, genomic and experimental analyses. By contrast, we show here, with a comparative analysis across 51 diverse species, that genes for extracellular proteins, which are likely to act as cooperative 'public goods', were not more likely to be carried on either: (1) plasmids compared to chromosomes; or (2) plasmids that transfer at higher rates. Our results were supported by theoretical modelling which showed that, while horizontal gene transfer can help cooperative genes initially invade a population, it has less influence on the longer-term maintenance of cooperation. Instead, we found that genes for extracellular proteins were more likely to be on plasmids when they coded for pathogenic virulence traits, in pathogenic bacteria with a broad host-range.

| S-EPMC7612097 | biostudies-literature

Broad host range plasmids can invade an unexpectedly diverse fraction of a soil bacterial community.

Project description:Conjugal plasmids can provide microbes with full complements of new genes and constitute potent vehicles for horizontal gene transfer. Conjugal plasmid transfer is deemed responsible for the rapid spread of antibiotic resistance among microbes. While broad host range plasmids are known to transfer to diverse hosts in pure culture, the extent of their ability to transfer in the complex bacterial communities present in most habitats has not been comprehensively studied. Here, we isolated and characterized transconjugants with a degree of sensitivity not previously realized to investigate the transfer range of IncP- and IncPromA-type broad host range plasmids from three proteobacterial donors to a soil bacterial community. We identified transfer to many different recipients belonging to 11 different bacterial phyla. The prevalence of transconjugants belonging to diverse Gram-positive Firmicutes and Actinobacteria suggests that inter-Gram plasmid transfer of IncP-1 and IncPromA-type plasmids is a frequent phenomenon. While the plasmid receiving fractions of the community were both plasmid- and donor- dependent, we identified a core super-permissive fraction that could take up different plasmids from diverse donor strains. This fraction, comprising 80% of the identified transconjugants, thus has the potential to dominate IncP- and IncPromA-type plasmid transfer in soil. Our results demonstrate that these broad host range plasmids have a hitherto unrecognized potential to transfer readily to very diverse bacteria and can, therefore, directly connect large proportions of the soil bacterial gene pool. This finding reinforces the evolutionary and medical significances of these plasmids.

| S-EPMC4817699 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data