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Sample GSM5135079 Query DataSets for GSM5135079
Status Public on Aug 08, 2021
Title MCF7 E2 siCTL YTHDC1 ChIP-Seq rep1
Sample type SRA
 
Source name MCF7 cell
Organism Homo sapiens
Characteristics cell type: human breast cancer cell line
treatment: E2 60 mins,siCTL
antibody: YTHDC1, CST,#77422
Treatment protocol In order to deprive hormone signaling in MCF7 cells, the cells were incubated in phenol red-free DMEM with 10% Charcoal-stripped fetal bovine serum (CS-FBS, Omega Scientific Inc) for 72 hours.EtOH or E2 were added to the cells for an hour.
Cells were labeled in media for 15 min with 700 ?M 4-thiouridine (4sU, Sigma-Aldrich) and harvested through centrifugation for 1 min at 1,000g.
Growth protocol MCF7 cells were incubated in DMEM media supplemented with 10% Fetal Bovine Serum (FBS, GenDEPOT) in 37°C CO2 incubator.
MCF7 cells were incubated in DMEM media supplemented with 10% Fetal Bovine Serum (FBS, GenDEPOT) in 37°C CO2 incubator. . In order to deprive hormone signaling in MCF7 cells, the cells were incubated in phenol red-free DMEM with 10% Charcoal-stripped fetal bovine serum (CS-FBS, Omega Scientific Inc) for 72 hours. To induce the estrogen signaling pathway, 100 nM 17-?-estradiol (E2) were added to the cells for an hour.
Extracted molecule genomic DNA
Extraction protocol Lysates were clarified from sonicated nuclei and chromatin were isolated with specific antibody.
DNA libraries were prepared using NEB Next Ultra II DNA library prep kit for illumina according to manufactorer's instructions
RNA was extracted using TRIzol (Life Technologies), 4sU-labeled RNA was exctracted according to Duffy et al., Mol Cell, 2015. M6A methylated RNA was immunoprecipitated by anti-m6A antibodies (202003, SYSY).
RNA libraries were prepared using NEB Next Ultra II RNA library prep kit for illumina according to manufactorer's instructions
 
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model NextSeq 550
 
Data processing All clean-reads were mapped to human reference genome hg19 by STAR v2.7.0, with parameters of –-genomeSAindexNbases 14 –outFilterMultimapNmax 10 –outFilterMismatchNmax 10.   Duplicated reads were further removed, and only unique aligned reads will be considered for later visualization and quantification.
hg19
bed
MINT-Seq, TT-Seq, MeRIP-Seq,RNA-Seq, ChIP-Seq raw data were de-multiplexed by bcl2fastq (v2.20), and quality-controlled with fastqc.
All clean-reads were mapped to human reference genome hg19 by STAR v2.7.0, with parameters of –-genomeSAindexNbases 14 –outFilterMultimapNmax 10 –outFilterMismatchNmax 10.   Duplicated reads were further removed, and only unique aligned reads will be considered for later visualization and quantification.
For gene/mRNA quantification, hg19 RefSeq gene annotation coordinates were used. And HOMER tool-sets were used to calculate the FPKM value of individual repeat elements, based on repeat mask annotation. 
The unique aligned reads were further converted to bigwig format with a fragment length of 75 on IGV browser.
Genome_build: hg19
Supplementary_files_format_and_content: bigWig
 
Submission date Mar 04, 2021
Last update date Aug 08, 2021
Contact name Ruoyu Wang
E-mail(s) ruoyu.wang@uth.tmc.edu
Organization name UTHealth McGovern Medical School
Street address 6431 Fannin St
City HOUSTON
State/province Texas
ZIP/Postal code 77054
Country USA
 
Platform ID GPL21697
Series (1)
GSE143441 N6-adenosine Methylation of Enhancer RNAs and YTHDC1 Facilitate Transcriptional Condensate Formation and 3D Chromatin Organization
Relations
BioSample SAMN18140430
SRA SRX10239459

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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