|
| Status |
Public on Dec 04, 2017 |
| Title |
F1-FiPSC-K27ac |
| Sample type |
SRA |
| |
|
| Source name |
F1-FiPSCs
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Induced pluripotent stem cells (iPSCs)
passage: 23-27
chip antibody: Histone H3K27ac antibody (pAb)
chip antibody vendor: Active Motif
chip antibody cat. #: 39133
|
| Treatment protocol |
Different human cell types: CiPSCs, EiPSCs, FiPSCs, CPCs, ECs, and FBs.
|
| Growth protocol |
Human induced pluripotent stem cells (iPSCs) were maintained in mTeSR-1 medium (STEMCELL Technologies) on Matrigel-coated plates. Endothelial cells (ECs) were maintained on 0.1% gelatin-coated plates with EGM-2 medium (Lonza). Cardiac progenitor cells (CPCs) were cultured on 0.1% gelatin-coated dishes in M199 (Gibco)/EGM-2 (3:1) medium supplemented with 10% FBS, 10 ng/ml basic fibroblast growth factor (bFGF, PeproTech), 5 ng/ml epithelial growth factor (EGF, PeproTech), 5 ng/mL insulin-like growth factor (IGF-1, PeproTech), and 5 ng/ml hepatocyte growth factor (HGF, PeproTech). Fibroblast cells (FBs) were cultured in DMEM medium (Gibco) supplemented with 10% FBS (Hyclone) and GlutaMax (Gibco).
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Total RNAs were extracted by RNeasy Plus Mini kit (Qiagen) according to the manufacturer’s protocol.
RNA-seq libraries were generated using the NuGEN Encore Complete RNA-seq Library Systems (Part No. 0311, NuGEN). Chromatin immunoprecipitation were performed using approximately 1x10^7 cells of human iPSCs, fibroblasts, endothelial cells, and cardiac progenitor cells. Cells were first cross-linked with 1% formaldehyde for 10 min at RT, and formaldehyde was quenched by glycine with a final concentration of 0.125 M. Chromatin was broken into small pieces with an average size of 0.5-2 kb using the Bioruptor (Diagenode). The sonicated chromatin was then incubated with 3-5 μg of primary antibodies overnight at 4°C. A small portion (10%) of chromatin without antibody incubation was kept as input DNA for each ChIP reaction. Subsequently, 75 μl of Dynabeads Protein A or Protein G were added and incubator for 4 h at 4°C with overhead shaking. Magnetic beads were then washed away and chromatin was eluted. Crosslink was then reversed and precipitated DNA was purified and resuspended in nuclease-free water. Sequencing libraries of immunoprecipitated DNA and input DNA were constructed according to an Illumina DNA library preparation protocol.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
F1-FiPSC-K27ac
processed data file:
ChIP-seq_enhancer_K27ac_vsd_exp.txt
ChIP-seq_promoter_K27ac_counts.txt
|
| Data processing |
ChIP-seq: bowtie -p 4 -q -m 1 -v 3 --sam --best --strata /data/bowtie_index/hg38
RNA-seq: hisat -x hg38 -1 R1.fq.gz -2 R2.fq.gz -S R.sam
Genome_build: hg38
Supplementary_files_format_and_content: Text files. ChIP-seq: Raw read counts for specific loci. RNA-seq: DESeq2 normalized log2 transformed reads.
|
| |
|
| Submission date |
Mar 24, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Joseph Wu |
| Organization name |
Stanford University School of Medicine
|
| Street address |
Stanford University School of Medicine
|
| City |
Stanford |
| State/province |
Carlifornia |
| ZIP/Postal code |
94305 |
| Country |
USA |
| |
|
| Platform ID |
GPL16791 |
| Series (1) |
| GSE97033 |
Cell Type-Specific Chromatin Signatures Underline Regulatory DNA Elements in Human Induced Pluripotent Stem Cells and Somatic Cells |
|
| Relations |
| BioSample |
SAMN06642837 |
| SRA |
SRX2671816 |
