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| Status |
Public on Sep 14, 2023 |
| Title |
ILC2, ATAC, rep3 |
| Sample type |
SRA |
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| Source name |
murine ILC2
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| Organism |
Mus musculus |
| Characteristics |
cell type: murine ILC2
strain: C57BL/6
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Human ILC2s were isolated from a TrimaAccel LRS Chamber containing peripheral blood of health donors with the RosetteSep Human ILC2 Enrichment Kit (Stem Cell). ILC2s and pro-inflammatory conditioned ILC2s (pcILC2s) were cultured at 2.25 x 105 cells/mL in aMEM medium supplemented with 20% FBS and 1% Pen-Strep along with 50 ng/ml rIL-7, IL-2, IL-25, IL-4 and rIL-33 for the ILC2s and 50 ng/ml rIL-7, 50 ng/ml rIL-12, IL-2, rIL-1b, and 25 ng/ml rIL-18. After 14-21 days, viability and purity was assessed by flow cytometry and nuclei were isolated by pelleting 25,000 - 200,000 in a fixed-angle centrifuge. Mouse ILC2s were cultured at 2.25 x 10^5 cells/mL for 6 days in complete media (RPMI-1640 supplemented with 10% FBS, 2 mM L-glutamine, 12 mM HEPES, 0.1 mM non-essential amino acids, 1 mM sodium pyruvate, 1% Pen/Strep, and 50 μM 2-mercaptoethanol) and supplemented with 10 ng/ml rIL-7 and rIL-33 (PeproTech), with the media changed every 2 days. For experiments in which cells were generated via cytokine-mediated skewing (pcILC2s), cells were cultured at 2.25 x 105 cells/mL for 48 hours in complete media supplemented with 10 ng/ml rIL-7 and rIL-33 (PeproTech). On Days 2 and 4, the media was replaced with complete R10 containing 10 ng/ml rIL-7, 10 ng/ml rIL-12, 10 ng/ml rIL-1b, 10 ng/ml rIL-15, 10 ng/ml rIL-2, and 5 ng/ml rIL-18.
For library preparation for both mouse and human ILC2s, cells were lysed prior to 30 minutes of transposition with an in house Tn5 transposase at 37°C. Immediately following transposition, samples were purified with Zymo Conc & Clean (Genesee) and stored at -20°C prior to library amplification. Fragments were amplified using 1× NEBnext PCR master mix (New England BioSciences) and custom Nextera PCR primers. Full libraries were amplified for five cycles, after which a test aliquot of each sample was taken to test 20 cycles to determine the additional number of cycles needed for the remaining 45 μL reaction. After the additional cycles were complete (average of 5-15 additional cycles), libraries were purified prior to a two-sided Ampure bead (Beckman Coulter) size selection to enrich for nucleosome-free fragments. Fragment size and concentration were determined by TapeStation 2000 and Qubit 4 (Agilent).
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| Library strategy |
ATAC-seq |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 4000 |
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| Description |
1x75 high output
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| Data processing |
Basecalls were performed using bcl2fastq
cutadapt (v. 1.12) was used to trim addaptor sequences. Reads were quality filtered using FASTX-ToolKit (v0.0.12) with paramters Q 33, -p 90, and q 20. Reads were aligned to mm10 mouse genome or hg38 human genome using STAR (v2.5.2b).
Peaks were called uing MACS v2.1.2 usingdefault parameters
Assembly: mm10 and hg38
Supplementary files format and content: bigWig files contain total read depth normalized signal, narrowPeak files contain MACS2 peak calls / DESeq2_peaks.
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| Submission date |
May 08, 2023 |
| Last update date |
May 30, 2024 |
| Contact name |
SONIA LAURIE |
| E-mail(s) |
sonia.laurie@gmail.com
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| Organization name |
UNC CHAPEL HILL
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| Street address |
125 Mason Farm Road
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| City |
CHAPEL HILL |
| State/province |
NC |
| ZIP/Postal code |
27514 |
| Country |
USA |
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| Platform ID |
GPL21103 |
| Series (2) |
| GSE231999 |
Multiomic Single Cell Evaluation Reveals Inflammatory Cytokines Affect Innate Lymphoid Cell Fate After Allogeneic Stem Cell Transplantation [ATAC-Seq] |
| GSE232003 |
Multiomic Single Cell Evaluation Reveals Inflammatory Cytokines Affect Innate Lymphoid Cell Fate After Allogeneic Stem Cell Transplantation |
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| Relations |
| BioSample |
SAMN35004305 |
| SRA |
SRX20260779 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM7307973_ILC2_ATAC_mouse_rep3_peaks.narrowPeak.gz |
4.9 Mb |
(ftp)(http) |
NARROWPEAK |
| GSM7307973_mILC2_ATAC_rep3_combined_STARAligned.out.sorted.shiftedExtended.scaled.openChromatin.bw |
249.8 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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