|
| Status |
Public on Mar 18, 2023 |
| Title |
ChIP_H3K4me3_dTAG-24h_mDPY30-dTAG_ES_Rep2 |
| Sample type |
SRA |
| |
|
| Source name |
mDPY30-dTAG E14TG2a
|
| Organism |
Mus musculus |
| Characteristics |
cell line: mDPY30-dTAG E14TG2a
cell type: embryonic stem cells
chip antibody: H3K4me3
|
| Treatment protocol |
Cells were treated with dTAG 13 for 6, 12,or 24 hours.
|
| Growth protocol |
mESCs were cultured in knockout DMEM (Gibco) with 15% fetal bovine serum (BVES500, Biovision), 1 mmol/L L-glutamine (Gibco), 1× Sodium pyruvate (Gibco), 1× NEAA (Gibco), 0.11 mmol/L 2-mercaptoethanol, 103 U/mL LIF (Millipore) and 1× penicillin/streptomycin (Gibco) at 37 °C and 5% CO2.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Lysates were clarified from sonicated cells and protein-DNA complexes were isolated with antibody.
Libraries were prepared according to Vazyme's instructions accompanying the VAHTS Universal DNA Library Prep Kit (Vazyme ND607-02). Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (32 to 52 exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with Illumina primers for 15 cycles and library fragments of ~250 bp (insert plus adaptor and PCR primer sequences) were band isolated from an agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Illumina NovaSeq 6000 following the manufacturer's protocols.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
The raw ChIP-Rx reads were trimmed by Trim Galore v0.6.6 (https://www.bioinformatics.babraham.ac.uk/projects/trim_galore/) and aligned to the human hg19 and mouse mm10 assemblies using Bowtie v2.3.5.1 with default parameters.
Low mapping quality reads (MAPQ < 30) and PCR duplicates were removed using SAMtools v1.9 and Picard v2.23.3 (https:// broadinstitute.github.io/picard/).
We then collected the spike-in read number for each of the ChIP-Rx samples with SAMtools v1.9 and generated the normalization factor as 1e6/spike-in_count.
The ENCODE blacklist regions were removed using bedtools v2.29.2.
Peak calling was performed by macs2 v2.2.6 with a q-value threshold of 0.05.
DiffBind R package v2.16.2 was used to identify differential binding peaks.
Assembly: mm10
Supplementary files format and content: bigWig
|
| |
|
| Submission date |
Jan 13, 2023 |
| Last update date |
Mar 19, 2023 |
| Contact name |
Aixia Song |
| Organization name |
Fudan University
|
| Street address |
Dong'an Road
|
| City |
Shanghai |
| ZIP/Postal code |
200032 |
| Country |
China |
| |
|
| Platform ID |
GPL24247 |
| Series (2) |
| GSE222845 |
H3K4me2/3 modulate the stability of RNA polymerase II pausing (ChIP-Seq) |
| GSE222848 |
H3K4me2/3 modulate the stability of RNA polymerase II pausing |
|
| Relations |
| BioSample |
SAMN32730281 |
| SRA |
SRX19028375 |
