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| Status |
Public on May 20, 2024 |
| Title |
CpG_2 |
| Sample type |
SRA |
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| Source name |
Bone marrow
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| Organism |
Mus musculus |
| Characteristics |
tissue: Bone marrow
cell line: Bone marrow derived macrophage
cell type: Macrophage
genotype: wt
treatment: stimulated with CpG
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Take out the cryovial and shake it in 37°C water bath to quickly thaw it within 1-2min.Cells were centrifugedfor 5min at 600×g at room temperature. Cell activity was detected with LUNA-FL TM and counted. CUT&Tag assay was performed as described previously (Kaya-Okur, Wu et al. 2019). Briefly, the cells are bound to ConcanavalinA-coated magnetic beads, and the cell membrane is permeabilized by Digitonin. The enzyme pA-Tn5 Transposaseprecisely binds the DNA sequence near the target protein under the antibody guidance and results in factor-targetedtagmentation.
DNA sequence is tagmented, with adapters added at the same time at both ends, which can be enrichedby PCR to form the sequencing-ready libraries. After the PCR reaction, libraries were purified with the AMPurebeads and library quality was assessed on the Agilent Bioanalyzer 2100 system.
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
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| Data processing |
library strategy: Cut & Tag
Raw data (raw reads) of fastq format were firstly processed using fastp (version 0.20.0). In this step, clean data (cleanreads) were obtained by removing reads containing adapter, reads containing ploy-N and low-quality reads fromrawdata. At the same time, Q20, Q30 and GC content of the clean data were calculated. All the downstreamanalyseswere based on the clean data.
Reference genome and gene annotation files were downloaded from genome website directly. Index of the referencegenome was built using BWA(v0.7.12) and clean reads were aligned to the reference genome using BWAmem. These reads were then filtered for high quality (MAPQ ≥ 13),we also removed reads that were not properlypairedand with PCR duplicates. Only uniquely mapped(MAPQ ≥ 13) and de-duplicated reads were used for further analysis.
All peak calling was performed with MACS2 (version 2.1.0) using ‘macs2 -q 0.05 -f AUTO--call-summits--nomodel --shift -100 --extsize 200 --keep-dup all’. By default, peaks with q-value threshold of 0.05 was usedfor all data sets
The position of peak summit around transcript start sites of genes can predict the interaction sites between proteinandgene. ChIPseeker (Yu et al., 2015) was used to retrieve the nearest genes around the peak and annotate genomic region of the peak. Peak-related genes can be confirmed by ChIPseeker, and then Gene Ontology(GO) enrichment analysis was performed to identify the function enrichment results. GO enrichment analysis wasimplemented by the GOseq R package, in which gene length bias was corrected. GO terms with corrected P-valueless than 0.05 were considered significantly enriched by peak-related genes. KEGG is a database resourcefor understanding high-level functions and utilities of the biological system, such as the cell, the organismandtheecosystem, from molecular-level information, especially large-scale molecular datasets generated by genomesequencing and other high-throughput experimental technologies (http://www.genome.jp/kegg/). We used KOBASsoftware to test the statistical enrichment of peak related genes in KEGG pathways.
Assembly: mm38
Supplementary files format and content: peak annotation of samples
Supplementary files format and content: tdf: Tiled Data File format used specifically in genomic data visualization. TDF files are created to efficiently store and display large genomic datasets, such as those generated by sequencing methods like CUT&Tag, in genome browsers.
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| Submission date |
May 14, 2024 |
| Last update date |
May 20, 2024 |
| Contact name |
Zhejun Cai |
| E-mail(s) |
caizhejun@zju.edu.cn
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| Organization name |
Zhejiang University School of Medicine
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| Department |
Department of Cardiology, The Second Affiliated Hospital
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| Street address |
88 Jiefang Road
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| City |
Hangzhou |
| State/province |
Zhejiang |
| ZIP/Postal code |
310009 |
| Country |
China |
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| Platform ID |
GPL24247 |
| Series (1) |
| GSE267400 |
Cut & Tag sequencing exhibited the chromosome binding regions of transcription factor IRF5 within murine bone marrow derived macrophages |
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| Relations |
| BioSample |
SAMN41390814 |
| SRA |
SRX24543192 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM8265209_CpG_2.tdf |
223.2 Mb |
(ftp)(http) |
TDF |
| GSM8265209_CpG_2_vs_CpG_a_IgG_1.peak_annotation.txt.gz |
13.5 Mb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
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