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Sample GSM3566928 Query DataSets for GSM3566928
Status Public on Jul 08, 2019
Title Wildtype ESC IP of H3K14ac
Sample type SRA
Source name WT_H3K14ac
Organism Mus musculus
Characteristics strain background: 12910la
cell line: J1
genotype/variation: Wildytpe
cell type: embryonic stem cells (ESCs)
chip antibody: H3K14ac (abcam; cat.# ab52946; lot# GR149741-26)
Treatment protocol The CDS region of Zscan4c was cloned into pCMV-FLAG vector.WT mESCs were transfected with 1000ng pCMV-tag2 vector. After transfection for 24h, the cells were treated with 500ug/ml G418 for 10 days and the medium were changed every 24h, then we picked monoclonal cells.
Growth protocol Mouse J1 embryonic stem cells (ESCs) were cultured under 5% CO2 at 37℃ on well plate coated with 0.2% gelatin (Sigma), in medium (Hyclone) that supplement with15% FBS (Hyclone), 10ng/ml mLIF (GeneScript), 1% Penicillin-Streptomycin (Solarbiol), 1% GlutaMAX(Solarbio), 1% nonessential amino acids(Gibco), 0.1 mM β-mercaptoethanol (Sigma).
Extracted molecule genomic DNA
Extraction protocol Cells were lysed and chromatin was extracted. Anti-H3K27ac, anti-H3K4me1 and anti-H3K27ac antibodies were used to immunoprecipiate FLAG-Zscan4 in mESCs. Proteins on chromatin associated with Zscan4 was digested and chromatin was decrosslinked at 65 degree overnight. DNA was extracted in phenol-chloroform.
For ChIP-seq, 5ng of Zscan4-assocated chromatin DNA was used in ChIP-seq library preparation. Libraries were prepared according to Illumina protocols and library fragments of ~250 bp were used for sequencing.
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model HiSeq X Ten
Data processing Illumina RTA software used for image identification and basecalling.
Adaptor sequence and low-quality sequence were trimmed by Cutadapt with parameters " -q 5 -m 30 -n 4 -O 1 -a AGATCGGAAGAGCACACGTCTGAACTCCAGTCAC -A AGATCGGAAGAGCGTCGTGTAGGGAAAGAGTGTAGATCTCGGTGGTCGCCGTATCATT", then sequence reads were mapped to mm10 genome using Bowtie2 with parameters "-5 1 --very-sensitive -p 20 -I 0 -X 1000 --no-mixed --no-discordant --un-conc-gz".
Enrichment bigwig file was generated by bamCompare from Deeptools with parameter "--scaleFactorsMethod readCount -e --minMappingQuality 1 --samFlagExclude 256 --pseudocount -1 --operation ratio"
Genome_build: mm10
Supplementary_files_format_and_content: bigwig file of histone enrichment
Submission date Jan 17, 2019
Last update date Jul 29, 2019
Contact name Weiyu Zhang
Phone 18902012072
Organization name Nankai University
Street address No.38 Tongyan Road, Jinnan District
City Tainjin
ZIP/Postal code 300350
Country China
Platform ID GPL21273
Series (1)
GSE125238 CHIP-seq analysis about Histones and Zscan4 after Zscan4 overexpression
BioSample SAMN10762835
SRA SRX5257675

Supplementary file Size Download File type/resource 264.0 Mb (ftp)(http) BW
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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