Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

MaTAR25 RNA Antisense Pulldown and Mass Spectrometry

ABSTRACT: It has been suggested that lncRNAs can interact with transcriptional regulators/co-factors to form ribonucleoprotein (RNP) complexes to regulate the expression of downstream genes in the cell nucleus. To identify potential interacting proteins of MaTAR25, we used two different paired sets of biotin-labeled antisense oligonucleotides targeting MaTAR25 for native RNA antisense oligonucleotide pull-down (RAP) in 4T1 cells followed by qRT-PCR to assess pull-down efficiency. Samples were also eluted from beads for mass spectrometry isobaric tags for relative and absolute quantitative (MS-iTRAQ) analysis to identify proteins that bind to MaTAR25, and PPIB as the corresponding control. We ranked the candidate interactors based on detectable peptides above background in both pair sets of oligonucleotide pull-downs, and selected candidates with at least 2-fold enrichment compared to corresponding PPIB oligo pull-down.

INSTRUMENT(S): Orbitrap Fusion Lumos, Q Exactive

ORGANISM(S): Mus Musculus (mouse)

TISSUE(S): Epithelial Cell, Cell Culture

DISEASE(S): Breast Cancer

SUBMITTER: Kung-Chi Chang

LAB HEAD: David L. Spector

PROVIDER: PXD017398 | Pride | 2020-12-23

REPOSITORIES: Pride

ACCESS DATA

Json Xml

Similar Datasets

Determination of the RNA linked to the paraspeckle in the GH4C1 cell line

Project description:The goal of this study was to determine the identity of the RNA linked to the paraspeckles in the rat GH4C1 cell line. GH4C1 cell were fixed with 1% paraformaldehyde in PBS. Then nuclei were purified, lysed and sonicated. RNA pull-down was performed using two antisense DNA biotinylated oligonucleotide probes that target the lncRNA Neat1. Streptavidin-magnetic beads were added to hybridization reaction and complexes were captured by magnets. RNA was isolated using NucleoSpinÂ®RNA XS (Macherey-Nagel).

2016-07-21 | E-GEOD-81972 | biostudies-arrayexpress

IP-MS:Protein precipitated by antisense (AS) DNA oligomers crossing the junction site of circNEIL3

Project description:To identify the circNEIL3 interacting protein, we performed RNA pull-down assays after transfecting into cells with biotin-labeled sense (S) or antisense (AS) DNA oligomers crossing the junction site of circNEIL3, and the pull-down proteins were subjected to SDS-PAGE and Coomassie blue staining. ~45 kDa Band pulled down by the AS probe was specifically excised , digested and used for mass spectrometry analysis.

2023-10-24 | PXD039394 | Pride

Identifying LINC00899 binding sites in HeLa cells with CHART-seq

Project description:LncRNAs represent a major transcriptional output of the human genome, but the function of many of these elements is unknown. For chromatin-localized lncRNAs, identification of genomic binding sites of these lncRNAs provides one opportunity to characterize their function. In this experiment, we have used capture hybridization analysis of RNA targets with high-throughput sequencing (CHART-seq) to identify the binding sites of the lncRNA LINC00899 in HeLa cells. LINC00899 is of interest as its depletion results in mitotic delay, suggesting a role in mitotic progression. This experiment contains 5 replicate batches where each batch contains a sample with antisense capture oligonucleotides (COs) to hybridize to and pull down the lncRNA transcript; an input control for the antisense pulldown; a control sample with sense COs, which should not hybridize to and pull down the lncRNA transcript; and an input control for the sense pulldown. All samples in the same batch were generated at the same time, and each pulldown (antisense or sense) was performed on chromatin obtained from independent cell cultures. All samples were subjected to high-throughput paired-end sequencing across two lanes.

2019-03-29 | E-MTAB-7418 | biostudies-arrayexpress

Mouse hippocampus RNA pull down LC-MS/MS

Project description:Antisense and sense probes of noncoding RNA GAS5 were used to perform the RNA pull down experiment on mouse hippocampus tissues. Then the proteins were identified by LC-MS/MS.

2021-09-09 | PXD023363 | Pride

Analysis of circ101093 binding proteins in A549 cells after RNA pull-down.

Project description:We used circular RNA 101093 and its antisense to perform RNA pull-down, and analyzed the peptides in blank group, antisense group and circular RNA 101093 group. The object of this group is to analyze circ101093-binding protein. Blank and antisense group are used as negative control group.

2022-05-20 | PXD025575 | Pride

Identification of proteins interacting with recombinant Arabidopsis Legumain-beta

Project description:Pull down assay with recombinant AtLEGbeta to identify interaction partners

2023-08-23 | PXD042260 | Pride

Identification of human gene LOC107984012 interacting proteins in AC16 cardiomyocytes by pull-down assays coupled to MS

Project description:In this study, pull down assays combined with mass spectrometry analysis were performed in AC16 cardiomyocytes transfected with control plasmid (OE-Vector) or LOC107984012-overexpression plasmid to identify the repertoire of LOC107984012 interacting proteins. Pierce™ Magnetic RNA-Protein Pull-Down Kit (Thermo Fisher) provides reagents to efficiently enrich RNA Binding Proteins. Sense or antisense of LOC107984012 were in vitro transcription using the T7 RNA transcription system (Large Scale RNA ProductionSystem-T7, Promega) and biotin-labeled with the Pierce™ RNA 3’ End Desthiobiotinylation Kit (Thermo Scientific). RNA was then purified with QIAquick PCR Purification Kit (Qiagen). 30 μg of whole-cell protein lysates were incubated with purified biotinylated transcripts for 1 h at 4°C with rotation, then they were eluted for mass spectrometry identification.

2024-01-26 | PXD037001 | Pride

DNMT1-associated long non-coding RNA regulate global gene expression and DNA methylation in colon cancer

Project description:The investigation of the association of long non-coding RNAs (lncRNAs) with DNMT1 by RIP-seq reveals that DNMT1 interacts with DACOR1. We identified the genomic occupancy sites of DACOR1 through ChIRP-seq, which we found to significantly overlap with known differentially methylated regions (DMRs) in colon tumors. Induction of DACOR1 in colon cancer cell lines significantly reduced their ability to form colonies in vitro, suggesting a growth suppressor function. Consistent with the observed phenotype, induction of DACOR1 led to the activation of tumor-suppressor pathways and attenuation of cancer-associated metabolic pathways. Notably, DACOR1 induction resulted in down-regulation of Cystathionine ?-synthase (CBS), which is known to lead to increased levels of S-adenosyl methionine (SAM) – the key methyl donor for DNA methylation. The DNA methyltransferase DNMT1 was immunoprecipitated and all associated RNAs were identified by RNA-seq and subsequent bioinformatic analyses. Samples were prepared for DNA sequencing through the ChIRP-seq protocol which uses antisense DNA oligos to bind to DACOR1 and pull down associated DNA in complex. We sequenced one sample with DACOR1 specific probes and non-specific DNA probes as control. Three samples were collected for each control lentivirus transduced V852 colon cancer line and DACOR1 lentivirus transduced V852. Samples were processed for RNA-sequencing and analyzed through differential expression analysis.

2015-09-15 | E-GEOD-58989 | biostudies-arrayexpress

Ago2-bound ShortRnas in HCT116 cell line and Dicer_hypomorphic(DICER_Ex5 cells) HCT116 cell line

Project description:DICER_Ex5 cells were created by disrupting exon 5 of the human Dicer gene using an AAV targeting construct, thereby interrupting a well conserved segment of the N-terminal helicase domain while sparing the RNase III domains. In DICER_Ex5 cells, Dicer is expressed at a level lower than the wild type. This experiment started with RIP-anti-Ago2 pull-down, followed by high throughput sequencing analysis in wild type and Dicer-hypomorphic HCT116 cell lines.

2015-05-17 | E-MTAB-2056 | biostudies-arrayexpress

Evidence for Antisense Transcription Associated with microRNAs in Arabidopsis

Project description:Exploring miRNA-related antisense transcription in Arabidopsis through RNA transcript profiling of smRNA pathway-defective mutants on a custom high-resolution oligonucleotide array. Two custom arrays were designed with 15,000 oligonucleotide (25-mer and 36-mer) probes in each array. The probes were selected tiling 200 base upstream and downstream (3 n.t. resolution) of miRNA target sites in Arabidopsis thaliana. The arrays were fabricated by Agilent Technologies, Santa Clara, CA. The 22 target genes on the arrays were chosen based on the abundance of associated smRNAs that mapped to the loci, the amplitude of the antisense transcription signals in published whole genome tiling microarray experiments and a representative cross section of miRNA families. For 15k arrays with 24 selected miRNA targets, a dye swap loop experiment design was utilized with 12 blocks for 7 genotypes on two chip arrays. The details of the experimental design are in Supplemental Table V of the manuscript. For the hen1-1 versus Ler-0 experiment, a dye swap with two versus three biological replicates and four array blocks was performed.

2009-04-10 | E-GEOD-15199 | biostudies-arrayexpress

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

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