Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

A technique to detect native R-loops genome-wide without immunoprecipitation

ABSTRACT: We adapted the nuclease-targeting approach CUT&RUN (Skene & Henikoff, eLife 2017) to map R-loops in vivo. We show that this approach accurately identifies R-loops that are sensitive to both Actinomycin D and RNase H and largely overlap with previously identified regions containing R-loops. We also show that MapR can identify R-loops genome-wide from much smaller input material than existing techniques.

ORGANISM(S): Homo sapiens

PROVIDER: GSE120637 | GEO | 2019/10/18

REPOSITORIES: GEO

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Similar Datasets

A technique to detect native R-loops genome-wide without immunoprecipitation

Project description:A technique to detect native R-loops genome-wide without immunoprecipitation

| PRJNA493813 | ENA

CUT&RUN-seq of H3K4me3, H3K27me3 and H3K27ac in mll2wt and mll2[Y2602A] mESC

Project description:CUT&RUN-seq of H3K4me3, H3K27me3 and H3K27ac in mouse ESCs carrying a homozygous point mutation in the catalytic domain of mll2[Y2602A]. CUT&RUN was performed according to Skene & Henikoff, 2017 and purified DNA was using for library preparation with NEBNext DNA Ultra II kit (NEB E7645S; input: 5 ng of DNA). Libraries were multiplexed and sequenced on a NextSeq500 (Paired-End; read length 40). Each sample is present in 3 biological replicates.

2023-11-15 | E-MTAB-13466 | biostudies-arrayexpress

Improved elution conditions for native co-immunoprecipitation.

Project description:BackgroundNative immunoprecipitation followed by protein A-mediated recovery of the immuno-complex is a powerful tool to study protein-protein interactions. A limitation of this technique is the concomitant recovery of large amounts of immunoglobulin, which interferes with down-stream applications such as mass spectrometric analysis and Western blotting. Here we report a detergent-based "soft" elution protocol that allows effective recovery of immunoprecipitated antigen and binding partners, yet avoids elution of the bulk of the immunoglobulin.Methodology/principal findingsWe assessed the performance of the soft elution protocol using immunoprecipitation of Adaptor protein complex 1 (AP-1) and associated proteins as a test case. Relative to conventional elution conditions, the novel protocol substantially improved the sensitivity of mass spectrometric identification of immunoprecipitated proteins from unfractionated solution digests. Averaging over three independent experiments, Mascot scores of identified AP-1 binding partners were increased by 39%. Conversely, the estimated amount of recovered immunoglobulin was reduced by 44%. We tested the protocol with five further antibodies derived from rabbit, mouse and goat. In each case we observed a significant reduction of co-eluting immunoglobulin.Conclusions/significanceThe soft elution protocol presented here shows superior performance compared to standard elution conditions for subsequent protein identification by mass spectrometry from solution digests. The method was developed for rabbit polyclonal antibodies, but also performed well with the tested goat and mouse antibodies. Hence we expect the soft elution protocol to be widely applicable.

| S-EPMC3063181 | biostudies-literature

Genome-wide mapping of H3K4me3 and H3K27ac in monocytes from SARS-CoV-2 convalescent patients.

Project description:Epigenetic alterations of monocytes from COVID-19 convalescent individuals and healthy controls have been analyzed by mapping trimethylated H3K4 (H3K4me3) and acetylated H3K27 (H3K27ac) genome-wide using CUT&RUN (Skene & Henikoff, 2017). Epigenetic reprogramming of myeloid cells represents a hallmark of trained innate immunity (Netea, Joosten et al., 2016). In general, active enhancers are marked by H3K27ac and transcription start sites are marked by H3K4me3 and H3K27ac (Kimura, 2013). A global analysis of histone modifications associated with transcription start sites (TSS) revealed a difference in the overall coverage of specific loci in SC-conv versus SC-naïve cells with SC-conv samples showing lower occupancy overall

2021-06-07 | GSE173568 | GEO

Sequential chromatin immunoprecipitation to detect SUMOylated MeCP2 in neurons.

Project description:The small ubiquitin-like modifier (SUMO) is a short peptide that can be covalently linked to proteins altering their function. SUMOylation is an essential post-translational modification (PTM). Because of its dynamic nature, low abundance levels, and technical limitations, the occupation of endogenous SUMOylated transcription factors at genomic loci is challenging to detect. The chromatin regulator Methyl CpG binding protein 2 (MeCP2) is subjected to PTMs including SUMO. Mutations in MeCP2 lead to Rett syndrome, a severe neurodevelopmental disorder. Here, we present an efficient method to perform sequential chromatin immunoprecipitation (Seq-ChIP) for detecting SUMOylated MeCP2 in neurons. This Seq-ChIP technique is a useful tool to determine the occupancy of SUMOylated transcription and chromatin factors at specific genomic regions.

| S-EPMC5600420 | biostudies-literature

Native RNA G quadruplex immunoprecipitation (rG4IP) from mammalian cells.

Project description:G quadruplex structures play an important role in regulating DNA replication and transcription and mRNA translation. Although there are several techniques that can determine its formation in vitro, the study of RNA G quadruplexes in vivo is not simple. In the current protocol, we describe an optimized technique (RNA G quadruplex immunoprecipitation [rG4IP]) to selectively pull down native cytoplasmic RNAs containing G quadruplex structures in mammalian cells. We also use a bicistronic plasmid to confirm and pinpoint the structure location. For complete details on the use and execution of this protocol, please refer to Surani et al. (2022).

| S-EPMC9127691 | biostudies-literature

Cell-free chromatin immunoprecipitation to detect molecular pathways in heart transplantation.

Project description:Existing monitoring approaches in heart transplantation lack the sensitivity to provide deep molecular assessments to guide management, or require endomyocardial biopsy, an invasive and blind procedure that lacks the precision to reliably obtain biopsy samples from diseased sites. This study examined plasma cell-free DNA chromatin immunoprecipitation sequencing (cfChIP-seq) as a noninvasive proxy to define molecular gene sets and sources of tissue injury in heart transplant patients. In healthy controls and in heart transplant patients, cfChIP-seq reliably detected housekeeping genes. cfChIP-seq identified differential gene signals of relevant immune and nonimmune molecular pathways that were predominantly down-regulated in immunosuppressed heart transplant patients compared with healthy controls. cfChIP-seq also identified cell-free DNA tissue sources. Compared with healthy controls, heart transplant patients demonstrated greater cell-free DNA from tissue types associated with heart transplant complications, including the heart, hematopoietic cells, lungs, liver, and vascular endothelium. cfChIP-seq may therefore be a reliable approach to profile dynamic assessments of molecular pathways and sources of tissue injury in heart transplant patients.

| S-EPMC10511822 | biostudies-literature

Ribosomal profiling in Cystic Fibrosis Bronchial Epithelial cells

Project description:We performed ribosome profiling in Cystic Fibrosis Bronchial Epithelial cells which data set is used to calcualte codon-specific ribosome occupancy using the approach described by Lareau et al. eLife 2014.

2017-05-18 | GSE74365 | GEO

Genomic Profiling of Native R-Loops with a DNA-RNA Hybrid Recognition Sensor Protein

Project description:R-Loops are unique RNA-containing chromatin structures, which participate in various key biological processes and associate with multiple human diseases. Accurately and comprehensively profiling R-Loops in the genome is crucial to study their functions under the physiological and pathological conditions. However, the existing methodologies have produced broad discrepancies in R-Loop profiling and an independent strategy is urgently needed. Here, we constructed an artificial DNA-RNA hybrid recognition sensor protein GST-His6-2XHBD with the hybrid-binding domain of RNase H1, and found GST-His6-2XHBD could specifically interact with DNA-RNA hybrids and behaves similarly to the anti-DNA-RNA hybrid S9.6 antibody in DRIPc-seq. Furthermore, we established a convenient method, R-loop CUT&Tag, by combination of GST-His6-2XHBD with a Tn5-based cleavage under targets and tagmentation approach. R-Loop CUT&Tag generates highly specific signals for native R-Loops, and can sensitively detect the R-Loop signals at the promoter, genebody and enhancer. R-Loop CUT&Tag also provides possibilities to genome-widely map the native R-Loops with limited materials, and may benefit the resolving of the broad discrepancies between multiple R-Loop mapping methods.

2021-02-22 | GSE156400 | GEO

Genome-wide mapping of native G-loops in living cells

Project description:G-quadruplex (G4) are four‑stranded DNA secondary structures that form in guanine‑rich regions of the genome, which can enhance or repress gene expression. An R-loop is a special triple-stranded nucleic acid structure formed when nascent RNA invades double-stranded DNA (dsDNA) during transcription. G-loops are constituted by one or more DNA G4 on one strand and a stable RNA/DNA hybrid on the other. We developed the HepG4-seq for mapping the native G4s and the HBD-seq for mapping native R-loops. We combined the HepG4-seq and HBD-seq to profile the genomic native G-loops, which are regions co-occupied by both native G4s and R-loops, in both HEK293 cells and mouse embryonic stem cells (mESCs).

2024-07-25 | GSE254764 | GEO

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