ChIP-seq studies of Th17 cells in Malat1 KO and WT mice
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE213014 | GEO | 2024/12/31
REPOSITORIES: GEO
Similar Datasets
Project description:Mouse embryonic fibroblasts (MEFs) with doxycycline (Dox)-inducible reprogramming cassette MKOS-ires-mOrange and a Nanog-GFP reporter were transduced with lentiviral Dox-inducible Ty1-BFP (blue fluoresce protein with Ty1 tag in the N-terminus) or Hic2-Ty1 (Hic2 with Ty1 tag in the C-terminus) expression vector with MOI 5. One day later reprogramming was initiated by the administration of Dox. 48 hours later, the cells for Hic2 ChIP-seq with anti-Ty1 antibody were crosslinked with 2mM of Disuccinimidyl Glutarate (DSG) for 45 min at room temperature (RT) under constant agitation, before being cross-linked with 1% formaldehyde solution for 10 min at room temperature. For KLF4 ChIP-seq, cells were only cross-linked with 1% formaldehyde solution for 10 min at room temperature. ChIP-seq library was generated with the NEBNext Ultra-II DNA Library Prep kit. Each library was uniquely barcoded using NEBNext Multiplex Oligos for Illumina. Samples were sequenced with the NextSeq High 40PE.
2024-04-26 | E-MTAB-13170 | biostudies-arrayexpress
Project description:Performing ChIP-seq analyses in clinical specimens has remained largely challenging due to multiple technical limitations and low quantities of starting material, resulting in low enrichments and poor signal-to-noise ratio. Here, we refined the original protocols for transcription factor ChIP-seq analyses in breast, prostate, and endometrial tumor tissue. In addition to the standard fixative formaldehyde, a second crosslinker Disuccinimidyl glutarate (DSG) was included in the procedure.
2018-12-27 | GSE114737 | GEO
Project description:We developed a general method based on RNA Antisense Purification (RAP) to identify the intermolecular RNA-RNA interactions of a target RNA (RAP-RNA). RAP-RNA identifies endogenous RNA-RNA complexes through in vivo crosslinking, RNA capture with antisense oligonucleotides, and high-throughput RNA sequencing. This approach provides a systematic view of other RNAs that interact with a target RNA, and furthermore can distinguish between direct and indirect RNA-RNA interactions through the use of crosslinking reagents with different reactivities with proteins and nucleic acids. We applied this method to numerous small and large noncoding RNAs, including U1 snRNA, Malat1 lncRNA, Xist lncRNA, U3 snoRNA, U17/Snora73a snoRNA and U12 snRNA. We examined the RNA and chromatin interactions of ncRNAs in mouse embryonic stem cells. We developed and applied three related protocols: RAP-RNA[AMT], RAP-RNA[FA], and RAP-RNA[FA-DSG]. In the RAP-RNA[AMT] protocol, we fixed direct RNA-RNA hybrids in mouse embryonic stem (ES) cells with 4'-aminomethyltrioxalen (AMT), a psoralen-derivative crosslinker; AMT generates inter-strand crosslinks between uridine bases in RNA but does not react with proteins. In the RAP-RNA[FA] protocol, we used a different crosslinking strategy to capture both direct and indirect RNA-RNA interactions: we fixed ES cells using formaldehyde (FA), which crosslinks protein-RNA and protein-protein interactions and thus can capture both indirect interactions as well as direct interactions that are caged or flanked by proteins. In the RAP-RNA[FA-DSG] protocol, we fixed with both FA and disuccinimidyl glutarate (DSG), a strong protein-protein crosslinker, to more efficiently capture RNAs linked indirectly through multiple protein intermediates.
2014-09-01 | E-GEOD-55914 | biostudies-arrayexpress
Project description:ChIP-seq analysis of Etv2, Brg1 and H3K27ac post Etv2 induction in MEF and ES/EB. Methods: EB or MEF cells were harvested as described above. ChIP assays for ETV2 and H3K27ac were performed from 2 x 10e6 MEFs or 1 x 10e7 EB cells, respectively. Briefly, cells were crosslinked with 1% formaldehyde for 10 min at room temperature and the reaction was quenched by glycine at a final concentration of 0.125 M. For the BRG1 ChIP-seq, cells were first crosslinked in 2 mM disuccinimidyl glutarate (DSG; Life Technologies: Cat. #20593) for 30 min then in 1% formaldehyde for 10 min, quenched with glycine for 5 min. The remainder of the ChIP protocol was performed following the protocol described by Magli et al19. We used antibodies against H3K27ac (Abcam, ab4729), ETV2 (Abcam, ab181847), and BRG1 (Abcam ab110641) for the respective pulldowns. For both of the pulldowns, protein A Dynabeads were added to the ChIP reactions and incubated for 30 minutes at room temperature. Magnetic beads were washed and chromatin was eluted. After crosslinking reversal, RNase A, and proteinase K treatment, ChIP DNA was extracted with the Min-Elute PCR purification kit (Qiagen). ChIP DNA was quantified with Quant-it PicoGreen dsDNA Assay Kit (Life Technologies). Sequencing libraries were prepared using equal amount of ChIP DNA per sample as per instructions of SMARTer® ThruPLEX® DNA-Seq Kit (Takara R400675) and SMARTer® DNA Unique Dual Index Kit - 24U Set A (Takara R400665). Sequencing was performed at the University of Minnesota Genome Center with the The NextSeq 550 high-throughput benchtop sequencer.
2022-01-12 | GSE168521 | GEO
Project description:The mitochondrial oxidative phosphorylation system (OXPHOS) contains subunits of dual genetic origin, namely the nuclear and the mitochondrial genome. Mounting the different subunits into functional OXPHOS complexes is aided by dedicated chaperones termed assembly factors. How unfunctional or superfluous proteins are recognized during assembly and directed to degradation is currently not understood. Here, we reveal that protein quality control is an integral part of assembly, which is organized in dedicated hubs gathering mitochondrial translation, protein import, assembly, and protein turnover. Fate-decision of newly synthesized proteins is achieved by molecular triaging engaging the evolutionary conserved prohibitin/m-AAA protease supercomplex and assembly chaperones. The localization of these competing activities in vicinity to the mitoribosomal tunnel exit allows for a rapid decision whether newly synthesized proteins are fed into OXPHOS assembly or to degradation. In total, three projects are hosted in this repository. 1) Phb1-Alfa DSG cross-linked immunoprecipitation 2) Metabolic switch from Glycerol to Glucose followed by Phb1-ALFA DSG cross-linked immunoprecipitation. 3) Phb2-ALFA DSG cross-linked immunoprecipitation.
2023-08-14 | PXD039717 | Pride
Project description:Even though the amine reactive BS2G and DSG cross-linkers have the same length of spacer and are based on N-hydroxysuccinimidic group, our data showed that each of them formed preferentially different cross-links. We demonstrated that the choice of cross-linker can have a significant impact on the output data for structural characterization of biomolecules. Using equimolar mixtures of DSG with d6-BS2G, and BS2G with d6-DSG, we established that the polar BS2G preferentially bound to polar regions of modified molecule, whereas non-polar DSG bound to hydrophobic regions. This phenomenon established that the mixture of polar and non-polar cross-linkers acted as an efficient tool for the determination of distance constraints in proteins.
2020-02-28 | PXD017299 | Pride
Project description:These data include the genome wide location analysis of Ogt by ChIP of Ogt in mouse ES cells. Precipitation of formaldehyde cross-linked chromatin prepared from mouse ES cell using specific antibody against OGT and IgG as control.
2013-01-06 | E-GEOD-39154 | biostudies-arrayexpress
Project description:We performed ChIP-seq in the GR18 cell line (derived from U2OS ATTC:HTB-96, stably transfected with an expression construct for GR), upon glucocorticoid (dexamethasone)treatment for 90 minutes. Cells were cross-linked with 1% formaldehyde for 3 minutes.
2019-03-11 | E-MTAB-7747 | biostudies-arrayexpress
Project description:ChIP-seq data from mouse adipocyte. Mature 3T3-L1 adipocytes were cross-linked with 1% formaldehyde 10 days after induction with MDI. Frozen cell pellets were submitted to the Broad Institute for subsequent analysis of Ebf1-bound regions using anti-Ebf1 antibody (Abnova H00001879-M01).
2014-06-10 | E-GEOD-58330 | biostudies-arrayexpress
Project description:Quantification of EGFR complex component from Formaldehyde cross-linked immuno-purification using PRM method
2018-08-08 | MSV000082800 | MassIVE