|
| Status |
Public on Mar 31, 2022 |
| Title |
H3K27me3_ODcells_si-IKBz_duplicate1 |
| Sample type |
SRA |
| |
|
| Source name |
odontoblasts like cells differentiated from dental pulp cells
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: dental pulp
antibody: anti-H3K27me3 antibody (ab192985)
differentiated cell type: odontoblasts like cells
sirna: si-IKBz
molecule subtype: ChIP-DNA
|
| Treatment protocol |
OD cells were forward-transfected with control siRNA (Negative Control DsiRNA: IDT) and siRNAs for human IκBζ (hs.Ri.NFKBIZ.13.2: IDT) at a final concentration of 10 nM using Lipofectamine RNAiMAX (Thermo Fisher Scientific) reagent and incubated for 24 h.
|
| Growth protocol |
OD cells were maintained in low glucose DMEM with ascorbic acid (100 g/ml) and -glecerophosphate (10 mM), LPS (0.1 mg/mll), and Dexamethasone (10-8 M))
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Lysates were clarified from sonicated nuclei and histone-DNA complexes were isolated with antibody.
Cells used for ChIP-seq analysis were fixed with formaldehyde and ChIP was performed using the SimpleChIP plus enzymatic chromatin IP kit and magnetic beads (Cell Signaling Technologies) following the manufacturer’s protocol. Equal amounts of chromatin (estimated as chromatin obtained from 4 x 106 cells) were used for precipitation of protein-DNA complex with ChIP-validated anti-H3K27ac antibody (ab4729: Abcam). After reverse-crosslinking with proteinase K treatment, chromatin was eluted using ChIP DNA clean and concentrator (Zymo Research). Libraries were sequenced using 150-bp paired-end reads on an NovaSeq instrument to a depth of >10 million mapped reads.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
Adapter trimming and quality check were conducted with Trim galore and then aligned to a reference genome (hg38) using Bowtie2 with `-seed` parameters to efficiently use multimap reads with assuring reproducibility. Tag directories were generated using `makeTagDirectory` command in HOMER and ChIP-seq peaks were identified using the `getDiffrentialPeaksReplicates.pl` command in HOMER, specifying `-style histone` to generate a high confidence set of peaks across duplicates. This HOMER command internally calls DESeq2 to calculate enrichment value for each peak using the individual raw counts and return only those peaks that pass 2-fold enrichment and a FDR-adjusted p-value of < 0.05. Input chromatin DNA was used as controls.
Genome_build: hg38
Supplementary_files_format_and_content: bigwig files were generated by bamCoverage commond.
|
| |
|
| Submission date |
Aug 06, 2021 |
| Last update date |
Mar 31, 2022 |
| Contact name |
Shigeki Suzuki |
| Organization name |
Tohoku University Graduate School of Dentistry
|
| Department |
Department of Periodontology and Endodontology
|
| Street address |
4-1, Seiryo-machi, Aoba-ku
|
| City |
Sendai |
| State/province |
Miyagi |
| ZIP/Postal code |
9808575 |
| Country |
Japan |
| |
|
| Platform ID |
GPL24676 |
| Series (1) |
| GSE181596 |
ChIP-seq analyses of human odontoblast-like (OD) cells transfected with siRNA for IKBz |
|
| Relations |
| BioSample |
SAMN20602856 |
| SRA |
SRX11663904 |
