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Genome-wide binding sites of AtDEK3 in Arabidopsis thaliana.

ABSTRACT: The scope of the project is to find DNA binding sites of the DNA binding protein AtDEK3. For the experiments AtDEK3 overexpressor (OE) plants have been obtained. The AtDEK3 was tagged with a CFP-tag. With ChIP the protein was pulled down and eluted DNA was send for sequencing. Gene ontology (GO) term classification of genes enriched for AtDEK3 binding indicated that AtDEK3 target genes are involved in diverse biological processes.

ORGANISM(S): Arabidopsis thaliana

PROVIDER: GSE55893 | GEO | 2014/11/14

SECONDARY ACCESSION(S): PRJNA241253

REPOSITORIES: GEO

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Genome-wide binding sites of AtDEK3 in Arabidopsis thaliana.

Project description:The scope of the project is to find DNA binding sites of the DNA binding protein AtDEK3. For the experiments AtDEK3 overexpressor (OE) plants have been obtained. The AtDEK3 was tagged with a CFP-tag. With ChIP the protein was pulled down and eluted DNA was send for sequencing. Gene ontology (GO) term classification of genes enriched for AtDEK3 binding indicated that AtDEK3 target genes are involved in diverse biological processes. ChIP Seq of three indenpendendt AtDEK3 OE and their respective inputs were used.

2014-11-14 | E-GEOD-55893 | biostudies-arrayexpress

RNA coimmunoprecipitations

Project description:Nascent polypeptides emerging from the ribosome during biogenesis can interact with many chaperones and protein homeostasis factors. The high sensitivity of RNA identification can be used to identify substrates of specific cotranslationally acting chaperones. Protein A-tagged (TAP-tag) chaperones associated with translating ribosomes were immunopurified by binding to magnetic IgG beads, washed, and chaperone complexes were eluted with TEV protease treatment. Immunopurified RNAs and total cell extract RNAs were isolated, reverse transcribed, coupled to Cy5 and Cy3 dyes, respectively, and comparatively hybridized to DNA microarrays

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An increase in the abundance of Tim23 in Arabidopsis thaliana results in the disruption of complex I and activation of mitochondrial biogenesis.

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2011-09-01 | E-MEXP-3309 | biostudies-arrayexpress

RBP9-TAP RIPseq in Trypanosoma brucei

Project description:RBP9-TAP was expressed in bloodstream T. brucei and bound mRNA was pulled down using IgG beads and eluted via TEV-protease cleavage. rRNA was depleted using RNAseH and rRNA hybridizing oligos.

2020-05-30 | E-MTAB-9093 | biostudies-arrayexpress

Mapping the differential distribution of the cohesin subunit Scc1 in Trypanosoma brucei brucei.

Project description:Chromatin immunoprecipitation of genomic loci in Trypanosoma brucei where Scc1 is deposited. This was achieved by deletion of one Scc1 allele and N-terminal tagging of the second Scc1 allele with a Ty1-tag. During the ChIP experiment, the DNA was crosslinked and cells were permeabilized with digitonin. DNA was fragmented by sonication with a Covaris instrument. DNA bound to Ty1-Scc1 was pulled down by using a BB2 anti-Ty1 antibody. Cross links are reversed and the DNA was purified and prepared for Illumina sequencing.

2018-11-06 | GSE119071 | GEO

DAXX genome binding/occupancy profiling by high throughput ChIP sequencing

Project description:Purpose: The goal of this study is to compare DAXX genome binding/occupancy by high throughput ChIP sequencing in control, wild type DAXX overexpressd cells (WT DAXX OE), and wild type DAXX SUMO interacting motif mutant overexpressd cells (DSM OE). Methods: The panel of MDA_MB-231 derived cell lines (control, WT DAXX OE, DSM OE) were cultured in complete DMEM. At about 90% confluency, the cells were crosslinked by adding 37 % formaldehyde to the final concentration of 1% for 10 min at room temperature. Crosslinking was stopped by adding glycine to the final concentration of 125 mM. Cells were lifted, washed with cold PBS, and pelleted by centrifugation. The cells were resuspended in a swelling buffer in the presence of the protease inhibitor cocktail (Sigma) and then pelleted and resuspended in the SDS lysis buffer. The lysates were transferred to a Covaris microTUBE and sonicated with an E220 Covaris Ultrasonicator. Chromatin fragmentation (~500 bps) was verified through agarose gel electrophoresis. The fragmented chromatins were diluted and incubated with a control IgG and the DAXX mAb (5G11) along with protein A/G magnetic beads. The beads were washed sequentially with a low salt buffer, high salt buffer, LiCl buffer, and TE buffer (twice). The immunoprecipitated chromatins were eluted at 65 °C for 15 min, and the eluted chromatins were subjected to proteinase K digestion at 65 °C for 3 h. The DNAs were recovered through a Qiagen mini-prep column. The immunoprecipitated DNAs were used for qPCR and library construction and high throughput sequencing using an Illumina Hi-Seq 2500 sequencer. Results: We surveyed genome-wide occupancy of DAXX using ChIP-seq technology. Overexpression of WT DAXX increased DAXX?s chromatin association specifically with lipogenic pathway genes where DSM OE reverse. Conclusions: Our study represents the first detailed analysis of DAXX occupancy in lipoegenic gene transciption.

2021-12-20 | GSE190783 | GEO

Transcriptome profiling of zebrafish neural crest mutants

Project description:Total RNA was extracted from morpholigcally abnormal and wildtype sibling larvae of neural crest mutants derived from ENU mutagenesis. The 3 end of fragmented RNA was pulled down using polyT oligos attached to magnetic beads, reverse transcribed, made into Illumina libraries and sequenced using Illumina HiSeq paired-end sequencing. Protocol: Total RNA was extracted from zebrafish embryos using Trizol and DNase treated. Chemically fragmented RNA was enriched for the 3 ends by pulled down using an anchored polyT oligo attached to magnetic beads. An RNA oligo comprising part of the Illumina adapter 2 was ligated to the 5 end of the captured RNA and the RNA was eluted from the beads. Reverse transcription was primed with an anchored polyT oligo with part of Illumina adapter 1 at the 5 end followed by 12 random bases, then an 8 base indexing tag, then CG and 14 T bases. An Illumina library with full adapter sequence was produced by 20 cycles of PCR. This data is part of a pre-publication release. For information on the proper use of pre-publication data shared by the Wellcome Trust Sanger Institute (including details of any publication moratoria), please see http://www.sanger.ac.uk/datasharing/

2014-04-28 | E-ERAD-243 | biostudies-arrayexpress

RNA coimmunoprecipitations

2013-04-05 | E-GEOD-45794 | biostudies-arrayexpress

Iron-deficiency-induced changes in wild type, ubc13A and cucumber CsUBC13 overexpressed Arabidopsis thaliana roots

Project description:CsUBC13 was identified via proteomics from iron starvation treated Cucumber root. ubc13A is an ABRC seed stock (CS51269). CS851269 was purchased from ABRC and confirmed as homozygous Atubc13A knock-out T-DNA mutant. We generated transgenic arabidopsis with ectopic expression of CsUBC13 gene under control of the cauliflower 35S promotor. Both genotypes and Col-0 were used to investigate the transcriptional response to Iron (Fe) deficiency. Wild type Col-0, ubc13A and transgenic overexpressor OE were grown under normal and iron-deficiency conditions. Roots were collected with 3 biological replicates.

2009-07-07 | E-GEOD-16964 | biostudies-arrayexpress

Maps of histone variant H3.V deposition in Trypanosoma brucei brucei

Project description:Chromatin immunoprecipitation of genomic loci in Trypanosoma brucei where histone variant H3.V is deposited. This was achieved by deletion of one H3.V allele and N-terminal tagging of the second H3.V allele with a Ty1 tag. During the ChIP experiment, the DNA was digested with MNase to obtain mononucleosomes. Nucleosomes containing H3.V were pulled down by using a BB2 anti-Ty1 antibody. Cross links are reversed and mononucleosomal DNA is purified and prepared for Illumina sequencing.

2018-08-24 | GSE118917 | GEO

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