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Status |
Public on Jan 14, 2023 |
Title |
E28 DZ, scATACseq |
Sample type |
SRA |
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Source name |
myotome
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Organism |
Sus scrofa |
Characteristics |
tissue: myotome developmental stage: embryonic day (E) 28 breed: DurocxTibetan pigs (DZ)
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Extracted molecule |
genomic DNA |
Extraction protocol |
Somite tissues of ZZ and DZ at E18, E21, and myotome tissues at E28 were isolated and transferred to RPMI1640 medium (GIBCO, 11875093) containing 10% Fetal Bovine Serum (HyClone, SH30070.03) on ice. Then, the tissues were washed with PBS three times and transferred to pre-warmed digestion medium containing 100 μg/mL DNase Ⅰ (Sigma-Aldrich, DN25) and 0.1 g/mL Collagenase I (Sigma, C2674) in RPMI1640. Tissues were shaken vigorously for 30 s and further incubated at 37°C for 30 to 40 minutes in an incubator with general shaking every 6 minutes to release cells. Released cells passed through a 70 μm cell strainer (BD, 352350) and were collected in 15 mL tubes. Sample viability was assessed via Trypan Blue (ThermoFisher) and an automatic cell counter (Countstar). Cells derived from different embryo ages and pigs were sorted and processed independently in all experiments. Thus, each cell can be traced back to the embryos and tissues it came from. scATAC-seq was performed using the 10× Chromium platform. All protocols to generate scATAC-seq data, including sample preparation, library preparation, and instrument and sequencing settings, are described below and are available at https://support.10xgenomics.com/single-cell-atac.
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Library strategy |
ATAC-seq |
Library source |
genomic single cell |
Library selection |
other |
Instrument model |
Illumina NovaSeq 6000 |
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Description |
10x Genomics
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Data processing |
Raw sequencing data aligned to the pig genome ATAC reference Sscrofa11 (10× reference database was prepared with gene annotation gff3 data v11.1.98 and genomic fasta data v11.1.98 downloaded from the Ensemble). The digital peaks matrix was quantified using the Cell Ranger ATAC (10× Genomics, v1.2.0) analysis pipeline. The following analysis was performed with the Seurat R package (v3.1.1) and Signac R package (v0.2.2). We kept only those valid barcodes that passed the following thresholds: (1) number of fragments ranging from 2000 to 30,000, (2) mitochondria ratio less than 2%, (3) transcription start site (TSS) enrichment score (by TSSEnrichment function) between 2 and 40, (4) strength of the nucleosome signal per cell (by NucleosomeSignal function) less than 5 and (5) fraction of reads overlapping peaks per cell greater than 50%. After this stringent quality control, we obtained 48,514 single cells. Normalization and latent semantic indexing (LSI) dimensionality reduction were performed for each sample independently with the RunTFIDF and RunSVD functions. Batch correction by samples was applied by Signac on the first 50 LSI components. We applied the UMAP and TSNE algorithms to the first 50 LSI components corrected by Signac. We identified 15 distinct clusters using the Seurat function FindClusters (resolution equal to 0.3) Supplementary files format and content: Sscrofa11 Supplementary files format and content: barcodes.tsv, peaks.bed, matrix.mtx
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Submission date |
Jun 24, 2022 |
Last update date |
Jan 14, 2023 |
Contact name |
Delin Mo |
E-mail(s) |
wangxy635@mail2.sysu.edu.cn
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Organization name |
Sun Yat-sen University
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Street address |
Xingang Road
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City |
Guangzhou |
ZIP/Postal code |
510006 |
Country |
China |
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Platform ID |
GPL26351 |
Series (2) |
GSE206913 |
Single-cell ATAC profiles of the somites and myotomes at embryonic stages E18, E21, and E28 from Tibetan pigs (ZZ) and Duroc x Tibetan pigs (DZ) |
GSE206914 |
Somites and myotomes at embryonic stages E18, E21, and E28 from Tibetan pigs (ZZ) and Duroc x Tibetan pigs (DZ) |
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Relations |
BioSample |
SAMN29406381 |
SRA |
SRX15890933 |
Supplementary file |
Size |
Download |
File type/resource |
GSM6266826_Y19SS167-barcodes.tsv.gz |
42.6 Kb |
(ftp)(http) |
TSV |
GSM6266826_Y19SS167-matrix.mtx.gz |
147.4 Mb |
(ftp)(http) |
MTX |
GSM6266826_Y19SS167-peaks.bed.gz |
1.0 Mb |
(ftp)(http) |
BED |
SRA Run Selector |
Raw data are available in SRA |
Processed data provided as supplementary file |
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