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Association of replication restart proteins with sites of increased transcriptional activity in Bacillus subtilis

ABSTRACT: Antibodies against DnaD and DnaB were used to immunoprecipitate genome bound complexes of these proteins in fast growing and slow growing cells as well as in two strains with a constitutively induced Pxis promoter or repressed Pxis promoter. CHIP-chip analysis was then carried out to determine their sites of interaction in vivo. CHI-chip analysis was carried out by Oxford Gene Technology (OGT) wwho also designed and prepared the arrays.

ORGANISM(S): Bacillus subtilis Bacillus subtilis subsp. subtilis str. 168

PROVIDER: GSE25736 | GEO | 2010/12/01

SECONDARY ACCESSION(S): PRJNA134083

REPOSITORIES: GEO

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Association of replication restart proteins with sites of increased transcriptional activity in Bacillus subtilis

Project description:Antibodies against DnaD and DnaB were used to immunoprecipitate genome bound complexes of these proteins in fast growing and slow growing cells as well as in two strains with a constitutively induced Pxis promoter or repressed Pxis promoter. CHIP-chip analysis was then carried out to determine their sites of interaction in vivo. CHI-chip analysis was carried out by Oxford Gene Technology (OGT) wwho also designed and prepared the arrays. Antibodies against DnaD and DnaB were used to immunoprecipitate genome bound complexes of these proteins in fast growing and slow growing cells, as well as in two strains with a constitutively induced Pxis promoter or repressed Pxis promoter. CHIP-chip microarray analysis was carried out by OGT who obtained the raw data. Data were visualized in CHIP Browser provided by OGT.

2010-12-01 | E-GEOD-25736 | biostudies-arrayexpress

Direct RNA sequencing enables single-nucleotide m6A detection in endogenous transcript isoforms

Project description:We report the direct RNA sequencing of HEK293 and a primary human mammary epithelial cell (HMEC) line using Oxford Nanopore based sequencing. Using this data, we built an algorithm to detect m6A modifications within the DRACH motif context. Evaluation of m6A sites was carried out with HEK METTL3 knockdown and HMEC ALKBH5 over expression cell lines.

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Genome-wide binding of replication initiation proteins in Bacillus subtilis

Project description:Initiation of DNA replication requires binding of the initiator protein, DnaA, to specific binding sites in the chromosomal origin of replication, oriC. In low G+C Gram-positive bacteria, the primosomal proteins DnaD and DnaB, in conjunction with loader ATPase DnaI, load the replicative helicase at oriC, and this depends on DnaA. DnaD and DnaB are also required to load the replicative helicase outside of oriC during replication restart, in a DnaA-independent manner. DnaA also binds to many sites around the chromosome, outside of oriC, and acts as a transcription factor at several of these. Using chromatin immunoprecipitation, we found that DnaD and DnaB, but not the replicative helicase, are associated with many of the chromosomal regions bound by DnaA in vivo in Bacillus subtilis. This association was dependent on DnaA and the order of recruitment was the same as that at oriC, but was independent of a functional oriC. The presence of DnaD and DnaB at the secondary (non-oriC) targets of DnaA in the absence of helicase loading indicates a possible role for DnaD and DnaB in modulating the activity of DnaA. The genome-wide binding profiles of DnaA, DnaD, DnaB and DnaC were determined. Binding profiles were determined in exponentially growing cells with and without HPUra treatment. Three biological replicates were analyzed per protein/treatment (one per array). Enrichment in immunoprecipitated samples versus total genomic DNA were determined.

2010-12-07 | E-GEOD-23686 | biostudies-arrayexpress

CHiCAGO: Robust Detection of DNA Looping Interactions in Capture Hi-C data

2016-05-18 | GSE81503 | GEO

ChIP-on-chip analysis of Ring1B, Ring1A, H2AK119u1 and H3K27me3 in mouse ES cells

Project description:ChIP-on-chip analysis was carried out to determine targets of Ring1B, Ring1A, H2AK119u1 and H3K27me3 in mouse ES cells. ChIP-on-chip analysis was carried out using the Mouse Promoter ChIP-on-chip Microarray Set (G4490A, Agilent, Palo Alto, Calif., USA). ESCs were subjected to ChIP assay using specific antibodies. Purified immunoprecipitated and input DNA was subjected to T7 RNA polymerase-based amplification as described previously (van Bakel et al., 2008). Labeling, hybridization and washing were carried out according to the Agilent mammalian ChIP-on-chip protocol (ver.9.0). Scanned images were quantified with Agilent Feature Extraction software under standard conditions.

2012-06-05 | E-GEOD-38504 | biostudies-arrayexpress

1f0e2786-a5a5-4414-a66b-00d73f46b6a6 - samples

Project description:Dataset contains four samples taken from a neonate with congenital KMT2A-rearranged Acute Lymphoblastic Leukemia patient (CHI-0391) with rare IKZF1 gene fusions. Sequencing was carried out using mRNA-seq sequencing on a Illumina NextSeq 500 machine

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Rybp ChIP-chip in mouse ES cells

Project description:Rybp binding in wild-type, Eed-KO, and Dnmt1-KO ES cells ChIP on chip analysis was carried out using the Mouse Promoter ChIP-on-chip Microarray Set (G4490A, Agilent-014716 and Agilent-014717, Palo Alto, Calif., USA). ESCs were subjected to ChIP assay using a Rybp antibody (Santa Cruz; H-115). Purified immunoprecipitated and input DNA was subjected to T7 RNA polymerase-based amplification as described previously (van Bakel et al., 2008). Labeling, hybridization and washing were carried out according to the Agilent mammalian ChIP-on-chip protocol (ver.9.0). Scanned images were quantified with Agilent Feature Extraction software under standard conditions.

2012-04-03 | E-GEOD-32289 | biostudies-arrayexpress

RNA-seq Analyzes Transcriptomes of dnaB-Ts and dnaB-Ts ΔahpC at Both Permissive and Non-permissive Temperature

Project description:To generate a bona fide model to study post-stress baterial programmed cell death (PCD), we used a temperature-sensitive E. coli mutant (dnaB-Ts) since temperature stress can be rapidly reversed while other stress, such as antibotic treatment, is difficult to be rapidly and completely stopped and removed for post-stress PCD analysis. When cells were shifted from permissive (30 °C) to non-permissive temperature (42 °C), the dnaB-Ts ΔahpC double mutant maintained full surival while survival of the dnaB-Ts single mutant dropped 3 orders of magnitude; similarly, the double mutant showed much higher survival during treatments with quinolones and β-lactams at permissive temperature. The purpose of the RNA-seq analyses is to uncover molecular mechanisms underlying the increased survival of the dnaB-Ts ΔahpC double mutant by comparing its transcriptional profiles with that of the dnaB-Ts single mutant at both permissive and non-permissive temperature. The results showed that multiple antioxidative systems and stress response pathways were highly upregulated due to a deficiency of ahpC when combined with a dnaB-Ts mutation. The high expression of these genes are consistent with the lower levels of toxic reactive oxygen species (ROS) in the dnaB-Ts ΔahpC mutant than in the dnaB-Ts single mutant. Thus, the RNA-seq data supported that ROS are an important lethal factor in bacterial suicide pathways when bacterial cells are exposed to harsh stress.

2018-05-12 | GSE114262 | GEO

CHiCAGO: Robust Detection of DNA Looping Interactions in Capture Hi-C data

Project description:Capture Hi-C (CHi-C) is a state-of-the art method for profiling chromosomal interactions involving targeted regions of interest (such as gene promoters) globally and at high resolution. Signal detection in CHi-C data involves a number of statistical challenges that are not observed when using other Hi-C-like techniques. We present a background model, and algorithms for normalisation and multiple testing that are specifically adapted to CHi-C experiments, in which many spatially dispersed regions are captured, such as in Promoter CHi-C. We implement these procedures in CHiCAGO (http://regulatorygenomicsgroup.org/chicago), an open-source package for robust interaction detection in CHi-C. We validate CHiCAGO by showing that promoter-interacting regions detected with this method are enriched for regulatory features and disease-associated SNPs. Three human CHi-C biological replicates were generated (comprising 1, 2and 3 technical replicates). Two mouse CHi-C biological replicates were generated (both comprising three technical replicates) and a mouse Hi-C dataset. The publicly available HiCUP pipeline (doi: 10.12688/f1000research.7334.1) was used to process the raw sequencing reads. This pipeline was used to map the read pairs against the mouse (mm9) and human (hg19) genomes, to filter experimental artefacts (such as circularized reads and re-ligations), and to remove duplicate reads. For the CHi-C data, the resulting BAM files were processed into CHiCAGO input files, retaining only those read pairs that mapped, at least on one end, to a captured bait. CHiCAGO then identified Hi-C restriction fragments interacting, with statistical significant, to captured baits.

2016-05-18 | E-GEOD-81503 | biostudies-arrayexpress

Immunoprecipitation Mass-Spectrometry (IP-MS) experiment reveals endogenous targets of ASFV pI73R

Project description:To characterize the cellular targets of the ASFV pI73R, we carried out an Immunoprecipitation Mass-Spectrometry (IP-MS) experiment. Whole-cell extracts prepared from PAM cells which infected with ASFV I73R-mHA were used to immunoprecipitate endogenous targets pI73R, with a matched IgG serving as a control. The immunoprecipitated proteins were then subjected to LC-MS/MS.

2021-11-29 | PXD030084 | iProX

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