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Sample GSM4728071 Query DataSets for GSM4728071
Status Public on Dec 07, 2020
Title LSC_scr_H3K27AC_rep1
Sample type SRA
 
Source name limbal stem cells
Organism Homo sapiens
Characteristics cell type: limbal stem cells
treatment: scramble treated
Treatment protocol For ChIP-seq, chromatin was fixed with 1% formaldehyde for 10 min and sheared in a sonification buffer. Fragmented DNA was incubated with primary antibodies overnight at 4℃ and then with Protein A/G Dynabeads for 1 h. The beads were collected and washed sequentially with the following solutions: high-salt buffer, low-salt wash buffer, and TE buffer. Elution of the immunoprecipitated protein/DNA complexes was performed using elution buffer at 65℃ for 4 h. Next, the protein/DNA complexes were incubated with proteinase K (Invitrogen) and RNase A (Invitrogen) for 1 h. De-crosslinked DNA was purified using the PCR Purification Kit (Qiagen). For ATAC-seq, In total, 50,000 cells were digested, collected, and lysed in ice-cold lysis buffer for 5 min. Then, Tn5 transposase reactions (Vazyme Biotech, TD501) were performed using the TruePrep DNA Library Prep Kit (Vazyme Biotech).
Growth protocol LSCs and SESCs were grown in 1:1 DMEM/F-12 medium containing 1% penicillin/streptomycin, 10% fetal bovine serum, 10 ng/ml EGF, 5 μg/ml insulin, 0.4 μg/ml hydrocortisone, 0.1 nM cholera toxin and 2 nM 3,3',5-triiodo-L-thyronine.
Extracted molecule genomic DNA
Extraction protocol Purified DNA was used to prepare ChIP-seq DNA libraries for Illumina sequencing by KAPA Hyper Prep Kit (KAPABIOSYSTEMS). For ATAC-seq library, the transposed DNA fragments were amplified and collected using the TruePrep DNA Library Prep Kit (Vazyme Biotech).
 
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model HiSeq X Ten
 
Data processing Reads were trimmed by trimmomatic tool and then aligned to human hg19 reference genome using BWA software (version 0.7.17).
Duplicated reads were removed by Picard Markduplicates (version 2.18.16) and only uniquely mapping reads were retained for further analysis.
MACS2 (version 2.1.1) was used to call peak. For sharp peaks, we used the following parameters: -f BAMPE -B --SPMR -q 0.001 --call-summits --fix-bimodal --seed 11521 --extsize 200. For broad peaks (H3K27me3 and H3K4me1), the following parameters were used: -f BAMPE -B --SPMR --fix-bimodal --extsize 500 --broad --broad-cutoff 0.1 --seed 11521.
Genome_build: hg19
Supplementary_files_format_and_content: Peak files with enriched genomic site and quantitative value.
 
Submission date Aug 14, 2020
Last update date Dec 07, 2020
Contact name Mingsen Li
E-mail(s) lims3@mail2.sysu.edu.cn
Organization name Sun Yat-sen University
Department Zhongshan Ophthalmic Center
Street address No. 7 Jinsui road Tianhe district
City Guanzhou
ZIP/Postal code 510060
Country China
 
Platform ID GPL20795
Series (2)
GSE156272 Core transcription regulatory circuitry orchestrates stratified epithelial homeostasis (ChIP-seq, ATAC-seq)
GSE156273 Core transcription regulatory circuitry orchestrates stratified epithelial homeostasis
Relations
BioSample SAMN15822621
SRA SRX8948970

Supplementary file Size Download File type/resource
GSM4728071_scr_H3K27AC_rep1_call_peak.xlsx 8.4 Mb (ftp)(http) XLSX
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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