Tnseq to identify recipient factor to modulate conjugative plasmid transfer
Ontology highlight
ABSTRACT:
INSTRUMENT(S): NextSeq 500
ORGANISM(S): Escherichia coli K-12
SUBMITTER:
PROVIDER: E-MTAB-11704 | biostudies-arrayexpress |
SECONDARY ACCESSION(S): E-MTAB-9713
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:Study of the possible existence of a replication fork trap in Vibrio cholerae. 1- FX85: EPV50(WT) grown in M9 minimal medium supplemented with 0.4 % fructose to exponential phase (0.2 OD 650 nm). 2- FX86: EPV50(WT) grown in M9 minimal medium supplemented with 0.4 % fructose to stationary phase (long overnight). 3- FX288: EGV140 (oriL3) grown in M9 minimal medium supplemented with 0.4 % fructose to exponential phase (0.2 OD 650 nm). 4- FX289:EGV140 (oriL3) grown in M9 minimal medium supplemented with 0.4 % fructose to stationary phase (long overnight). 5- FX290: EGV111 (oriR4) grown in M9 minimal medium supplemented with 0.4 % fructose to exponential phase (0.2 OD 650 nm). 6- FX291:EGV111 (oriR4) grown in M9 minimal medium supplemented with 0.4 % fructose to stationary phase (long overnight). 7- FX355:EPV50 (WT) grown in LB medium to exponential phase (0.2 OD 650nm) 8- FX356:EPV50 (WT) grown in LB medium to stationary phase (overnight) 9- FX286:EGV140 (oriL3) grown in LB medium to exponential phase (0.2 OD 650nm) 10- FX287:EGV140 (oriL3) grown in LB medium to stationary phase (overnight) 11- FX292:EGV111 (oriR4) grown in LB medium to exponential phase (0.2 OD 650nm) 12- FX49: MCH1 (WT monochromosome) grown in M9 minimal medium supplemented with 0.4 % fructose to exponential phase (0.2 OD 650 nm). 13- FX48: MCH1 (WT monochromosome) grown in M9 minimal medium supplemented with 0.4 % fructose to stationary phase (long overnight). 14- FX11: EGV369 (oriL3 monochromosome) grown in M9 minimal medium supplemented with 0.4 % fructose to exponential phase (0.2 OD 650 nm). 15- FX12: EGV366 (oriR4 monochromosome) grown in M9 minimal medium supplemented with 0.4 % fructose to exponential phase (0.2 OD 650 nm). 16- FX296 EPV50 M9 Exp 17- FX294 EPV50 M9 Stat 18-FX316 EGV140 M9 Exp 19- FX315 EGV111 M9 Exp 20-FX318 MCH1 M9 Exp 21- FX317 MCH1 M9 Stat 22- FX320 EGV369 M9 Exp 23- FX319 EGV366 M9 Exp Chromosomal DNA was extracted using the Sigma GenElute bacterial genomic DNA kit. 5 μg of DNA were used to generate a genomic library according to Illumina's protocol. The libraries and the sequencing were performed by the High-throughput Sequencing facility of the I2BC (http://www.i2bc.paris-saclay.fr/spip.php?article399〈=en,CNRS, Gif-sur-Yvette, France). Genomic DNA libraries were made with the ‘Nextera DNA library preparation kit’ (Illumina) following the manufacturer’s recommendations. Library quality was assessed on an Agilent Bioanalyzer 2100, using an Agilent High Sensitivity DNA Kit (Agilent technologies). Libraries were pooled in equimolar proportions. 75 bp single reads were generated on an Illumina MiSeq instrument, using a MiSeq Reagent kit V2 (500 cycles) (Illumina), with an expected depth of 217X. Reads were aligned on the in silico reconstituted genome of the cognate strain using BWA software. An in-lab written MATLAB-based script was used to perform marker frequency analysis. Data were normalized by dividing uniquely mapping sequence reads by the total number of reads. Enrichment of uniquely mapping sequence reads in 1 kb non-overlapping windows were calculated and plotted against the chromosomal coordinates.
2021-05-07 | E-MTAB-7193 | biostudies-arrayexpress
Project description:In order to dissect the nuclear RNA quality control (QC) mechanisms, we used a powerful assay based on global perturbation of mRNP biogenesis in yeast Saccharomyces cerevisiae by the bacterial Rho helicase. We have monitored the global RNA levels upon Rho activation and compare it to basal RNA level in WT strain, which have revealed down-regulated transcripts. Then, the same experiment was performed but in a strain lacking Rrp47, giving us a precise overview of the QC targets. We also followed some QC components (Nrd1, Nab3, Trf4 and Rrp6) along the yeast chromosomes by ChIP-seq before and after perturbation of mRNP biogenesis by Rho (crosslinked dataset).
2019-12-04 | E-MTAB-7538 | biostudies-arrayexpress
Project description:We analyzed gene expression during conjugative transfer of plasmid RP4. Pairs of rifampicin-susceptible (RifS) and -resistance (RifR) strains of Pseudomonas putida KT2440 were conjugated for 10 minute on filter membrane in the presence of rifampicin to discriminate the expression changes in the donor and recipient cells.
2020-03-13 | GSE146879 | GEO
Project description:Integrative and conjugative elements (ICEs), a.k.a., conjugative transposons, are mobile genetic elements involved in many biological processes, including the spread of antibiotic resistance. Unlike conjugative plasmids that are extra-chromosomal and replicate autonomously, ICEs are integrated in the chromosome and replicate passively during chromosomal replication. It is generally thought that ICEs do not replicate autonomously. We found that when induced, Bacillus subtilis ICEBs1 replicates as a plasmid. The ICEBs1 origin of transfer (oriT) served as the origin of replication and the conjugal DNA relaxase served as the replication initiation protein. Autonomous replication of ICEBs1 conferred genetic stability to the excised element, but was not required for mating. The B. subtilis helicase PcrA that mediates unwinding and replication of Gram-positive rolling circle replicating plasmids was required for ICEBs1 replication and mating. Nicking of oriT by the relaxase and unwinding by PcrA likely directs transfer of a single-strand of ICEBs1 into recipient cells. This SuperSeries is composed of the SubSeries listed below.
2009-10-16 | GSE18380 | GEO
Project description:To perform Tn-seq experiments random distribution of the transposon in the genome of the recipient cells is essential. In case transposon is delivered into the host cell by conjugation, donor may affect the distribution of the transposon in the genome of the recipient cell. Here, we provide two Tn-seq performed with two different donor strains. FX371 corresponds the Tn-seq using E. coli B-2163 as donor strain. FX442 corresponds to the Tn-seq using E. coli MFDpir as donor strain.
2020-11-12 | E-MTAB-9713 | biostudies-arrayexpress
Project description:Bacillus velezensis strain GH1-13 with a native conjugative plasmid (pBV71) is thought to be beneficial to the bacterium, although no information on its effects exists. Here we show that strain GH1-13 frequently lost the plasmid during normal growth conditions in a rich medium and changed the morphology and sensitivity to selenite and tellurite. Compared to the plasmid-cured cells, the wild-type and complemented cells exhibited multicellular behavior with the expression of conjugative type IV pili and regulatory Rap homologous genes that regulate the interconnection between conjugation and biofilm formation. Further omics-based analyses of morphogenesis, biofilm formation, and antibiotic synthesis suggest that the conjugative plasmid activates envelope stress responses in association with increased biosynthesis of extracellular polysaccharide and antibiotics for protective functions of the host during exponential phase.
2021-05-17 | PXD017305 | Pride
Project description:Aim: to identify the transcriptional responses to acquisition of conjugative megaplasmids by Pseudomonas fluorescens, and how these effects are impacted by known compensatory mutations. Full abstract TBC.
2021-04-17 | GSE151570 | GEO
Project description:Pervasive Transcription originates from spurious promoters present in both orientations within coding sequences. Spurious transcripts are typically untranslated and thus for the most part are terminated by transcription termination factor Rho. They can be studied by impairing Rho activity, which in this study was achieved by depleting cells for Rho co-factor NusG. To this end, strains in which the only chromosomal copy of the nusG gene is transcribed from a promoter that can be repressed by arabinose (ARA) were used. This study was aimed at determining how pervasive transcription affect H-NS mediated gene silencing in Salmonella Pathogenicity Island 1 (SPI-1). In particular, the analysis concentrated on the region surrounding the master regulator gene HilD. The activities of promoters in this region were assessed performing semiquantitative 5’RACE-Seq. RNA extracted from cells exposed to the indicated treatments was reverse-transcribed with gene-specific primers in the presence of a template-switching oligonucleotide (TSO). The latter carries three 3’-terminal riboguanosines that hybridize to the polycytosine overhang (typically 3 or 4 Cs) added by the RT enzyme when it reaches the 5’ end of the RNA.This allows the RT enzyme to switch template and polymerise the TSO. Subsequent PCR amplification, with a primer complementary to the TSO and another annealing inside the reverse transcribed region, followed by sequence analysis, allows identifying the 5’ end of the RNA as the sequence lying immediately adjacent to the poly-G track. Performing the PCR step semiquantitatively provides information as to relative RNA levels. Here, the technique was adapted to high throughput sequencing by using PCR primers carrying Illumina adapters at their 5’ ends. Three strains were used, all three carrying the ARA-repressible nusG allele, MA12996, MA14302 and MA14358. This last strain carries a deleted version of SPI-1 with the Anhydrotetracycline (AHTc)-inducible Ptet promoter positioned at edge of the H-NS patch covering hilD, which made it possible to study the effects of NusG depletion in the presence or absence of overlapping transcription.
2022-07-04 | E-MTAB-11419 | biostudies-arrayexpress
Project description:we examined the three different mature biofilms and searched the genes which promoted the rapid biofilm formation when their population hosing the plasmids. We investigate the global transcriptional differences between the non-conjugative or conjugative plasmid-carrying and plasmid-free strains.
2007-05-15 | E-MEXP-953 | biostudies-arrayexpress
Project description:Rhizobia are gram-negative bacteria able to establish a symbiotic interaction with leguminous plants. Due to their nitrogen fixing capacity, the study of these microorganisms has acquired great relevance for the agriculture. Rhizobia usually harbor many plasmids in their genome which can be transferred to other organisms by conjugation. Two main mechanisms of regulation of rhizobial plasmid transfer have been described: Quorum sensing (QS) and rctA/rctB system. Nevertheless, new genes and molecules that modulate conjugative transfer have been recently described, demonstrating that new actors can tightly regulate the process. In this work, by means of bioinformatics tools and molecular biology approaches, two hypothetical genes are identified as playing key roles in conjugative transfer. These genes are located between conjugative genes of plasmid pLPU83a from Rhizobium favelukesii LPU83, a plasmid that showed a conjugative transfer behavior depending on the genomic background. One of the two mentioned genes, rcgA, is essential for conjugation, while the other, rcgR, acts as an inhibitor of the process. In addition to introducing this new regulatory mechanism, we show evidence of the functions of these genes in different genomic backgrounds, and confirmed that homologous proteins from non-closely related organisms play the same function. These findings set up a cornerstone for a new molecular circuit of conjugative transfer of plasmids.
2022-08-29 | PXD033583 | Pride