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| Status |
Public on Oct 06, 2023 |
| Title |
WT_IgECLIL33_ATAC rep1 |
| Sample type |
SRA |
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| Source name |
HSMCs
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| Organism |
Homo sapiens |
| Characteristics |
cells: Human skin mast cells
tissue: Skin biopsy
treatment: IL33, IgE anti-NP and NP-HSA
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| Treatment protocol |
IgE receptor cross-linking (IgECL) on HSMCs was accomplished through sensitization with 1 μg/mL chimeric human IgE anti-NP antibody (MCA333S, BIO-RAD, Hercules, CA) for 24 hours. The sensitized hSMCs were challenged with 100 ng/mL NP-HSA (N-5059, LGC, Petaluma, CA) for 2 additional hours. For single or combined cytokine and IgECL treatments, IL-33 (50 ng/mL, 200-33, PEPROTECH, Cranbury, NJ) was added to hSMCs for a total of 26 hours.
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| Growth protocol |
Cells enriched by Percoll density-dependent sedimentation were adjusted to a concentration of 1 × 106 cells/ml in serum-free X-VIVOTM 15 medium (Lonza BioWhittaker) containing 100 ng/ml recombinant human SCF (Peprotech) in 24-well plates. The culture medium was changed weekly and wells were split when cell concentration doubled. The percentages of mast cells were assessed cytochemical by metachromatic staining with toluidine blue. Mature mast cells at ~ 100% purity and viability are obtained by 6-8 wk of culture, and 8- to 16-wk-old mast cells were used in these experiments.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Briefly, 50,000 HSMCs that were either untreated or treated with IgE receptor crosslinking were spin down and washed with cold PBS once. Then the cells were resuspended in 50 uL cold ATAC-RSB-lysis buffer (10mM Tris-HCl pH 7.4, 10mM NaCl, 3mM MgCl2, 0.1% NP-40, 0.1% Tween-20 and 0.01% Digitonin) and incubated for 3 minutes. The lysis buffer was immediately washed out with 1 mL ATAC-RSB buffer (10mM Tris-HCl pH 7.4, 10mM NaCl, 3mM MgCl2 and 0.1% Tween-20). The cell pellet was resuspended in 50 ul transposition mixture (25 uL 2X TD buffer, 2.5 uL transposase (Illumina, FC-121-1030), 16.5 uL PBS, 0.5 uL 1% digitonin, 0.5 uL 10% Tween-20, 5 uL H2O) and incubated at 37 C for 30 minutes. The reaction was stopped by adding 2.5 uL of 0.5M EDTA pH 8 and transposed DNA was purified using Qiagen MiniElute PCR purification kit (Qiagen).
Purified DNA was amplified using the following condition: 72℃ for 5 min, 98 ℃ for 30 s, and 7 cycles: 98 ℃ for 10 s, 63 ℃ for 30 s, 72 ℃ for 1 min. The amplified libraries were cleaned up, size-selected, and the quality and quantity of libraries were assessed on 4150 TapeStation System (Agilent, CA).
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| Library strategy |
ATAC-seq |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
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| Data processing |
Raw sequencing reads (average 80 million paired-end reads, 2 biological replicates for each treatment) were aligned to the hg38 reference genome using Bowtie2 with very-sensitive and -x 2000 parameters.
The read alignments were filtered using SAMtools to remove mitochondrial genome and PCR duplicates.
Peaks were identified by MACS2 with the q-value cut-off of 0.05
Genome_build: hg38
Supplementary_files_format_and_content: Tdf and bedgraph files for each sample
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| Submission date |
Feb 16, 2022 |
| Last update date |
Oct 06, 2023 |
| Contact name |
HUA HUANG |
| E-mail(s) |
huangh@njhealth.org
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| Phone |
3033981281
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| Organization name |
National Jewish Health
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| Department |
Immunology and Genomic Medicine
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| Lab |
Huang lab
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| Street address |
1400 Jackson street, K511
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| City |
Denver |
| State/province |
Colorado |
| ZIP/Postal code |
80206 |
| Country |
USA |
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| Platform ID |
GPL24676 |
| Series (2) |
| GSE196883 |
IL-33 priming and antigenic stimulation synergistically promote the transcription of proinflammatory cytokine and chemokine genes in human skin mast cells [ATAC-Seq] |
| GSE196895 |
IL-33 priming and antigenic stimulation synergistically promote the transcription of proinflammatory cytokine and chemokine genes in human skin mast cells |
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| Relations |
| BioSample |
SAMN25997966 |
| SRA |
SRX14204810 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM5903603_ATAC_HSMCs_IgEIL33_1_treat_pileup.bdg.tdf |
463.3 Mb |
(ftp)(http) |
TDF |
| GSM5903603_ATAC_HSMCs_IgEIL33_1_treat_pileup.bedGraph.gz |
935.5 Mb |
(ftp)(http) |
BEDGRAPH |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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