|Public on Jan 06, 2021
|Primordial hemogenic endothelium from H9
|cell type: Inducible SOX17 H9 ESC
description: Total cell at D4
|Inducible SOX17 H9 ESC s were differentiated in ColIV coating plate. Singular Cells were plated 5,000 cells/cm2 to 10cm plates coated with ColIV (Sigma-Aldrich). After 1 day, media was changed to IF9S media with 50 ng/ml FGF, 2 mM LiCl, 15 ng/ml Activin A, and 50 ng/ml BMP4. On day 2, media was changed to IF9S media with 50 ng/ml VEGF, 2.5 uM SB431542 and 50 ng/ml FGF and D4 PHE cells were collected (1 X 106 cells).
|Sample followed the Millipore EZ-Magna CHIP A/G protocol. Eluted CHIP DNA IN 50 uL of elution buffer from kit. Lysates were clarified from sonicated nuclei and SOX17-DNA complexes were isolated with antibody.
DNA libraries were prepared for sequencing using standard Illumina protocols
|Illumina HiSeq 3000
|ATAC-seq fragments were mapped to human genome by BWA with default settings. Human genome were divided into 32 bp bins and the number of reads in each bin was counted. In order to smooth the data, reads were extended to 200 bp. To normalize signals across ATAC-seq datasets, the number of reads in each dataset was reduced by random sampling to the smallest number of reads present in the datasets. ATAC-seq peaks were called by MACS2.
ChIP-seq fragments were aligned by BWA (version 0.7.15) with quality threshold at 5 for read trimming and all the other options in default settings. Normalized SOX17 ChIP-seq signals were calculated by MACS2 by using all the tags at the same loci.
RNA-seq fragments were aligned by STAR (version 2.5.2b) to human genome with gene annotations from GENCODE (version 27). Transcript expression levels were quantified by RSEM (version 1.3.0).
Supplementary_files_format_and_content: ATAC-seq BED files contain peaks called by MACS2
Supplementary_files_format_and_content: ATAC-seq bigWig files contain normalized ATAC-seq signals as described in ATAC-seq data processing step
Supplementary_files_format_and_content: ChIP-seq bigWig files contain normalized SOX17 fold enrichment over IgG control calculated by MACS2
Supplementary_files_format_and_content: RNA-seq TSV files contain gene expression levels quantified by RSEM
|Nov 13, 2019
|Last update date
|Jan 06, 2021
|Igor I. Slukvin
|University of Wisconsin
|1220 Capitol Court
|Profiling of melocular mechanism of SOX17 effect during hematopoieisis of human ES cells