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| Status |
Public on May 01, 2024 |
| Title |
Mouse_E10.5_PA2_GFP |
| Sample type |
SRA |
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| Source name |
pharygneal arch 2
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| Organism |
Mus musculus |
| Characteristics |
tissue: pharygneal arch 2
age: E10.5
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| Extracted molecule |
total RNA |
| Extraction protocol |
Micro-dissected forming pinna prominences from Hmx1wt/EGFP embryos were enzymatically dissociated using 0.5% trypsin/1× EDTA for 10 minutes at 37 °C (for E10.5 and E12.5 stages) and papain digestion mix for 7 minutes (for E14.5 stage). The treated tissues were then rinsed in ice-cold 1× DMEM, filtered, and GFP+ cells were enriched using FACS (Sony, MA900). These collected cells were resuspended and diluted in 1× DPBS (Cytiva) to a concentration of 1×106 cells per ml
Gene expression libraries were prepared using GEXSCOPE Single Cell RNA Library Kits v.2 (Singleron, catalog no. SD-4180022). The single-cell suspensions were loaded onto the GEXSCOPE microchip, which facilitates automated single cell capture, cell lysis, cellular mRNA capture, and molecular labeling via the Singleron Matrix instrument. The barcoded cDNA was amplified and utilized for constructing single cell NGS libraries. The quality and quantity of the resulting libraries were assessed using Qubit 4.0 (Thermo Fisher Scientific) and Qseq100 (Bioptic)
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic single cell |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
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| Data processing |
The scRNA-seq reads were processed using the CeleScope pipeline, details of which are available at https://github.com/singleron-RD/CeleScope. This processing generated a 10x Genomics-like raw feature-barcode matrix suitable for downstream filtering and data integration using Seurat (version 4.3.0). For cell filtering, we assumed that both the log10-transformed gene counts and UMI counts per cell for each sample followed a normal distribution. We calculated the mean ± 1.96 times the standard deviation, defining the 95% confidence interval, to determine the lower and upper thresholds. These thresholds were then converted back to the original scale (as the power of 10 of the log values) to establish criteria for excluding low-quality cells. Furthermore, cells exhibiting a mitochondrial gene expression ratio exceeding 5% were also categorized as low quality.
Data integration was accomplished using the CCA2-based pipeline of Seurat package. The SelectIntegrationFeatures function was utilized to identify variable features across datasets for integration. Subsequently, the FindIntegrationAnchors function was employed to identify anchor pairs between datasets, followed by the IntegrateData function to create an integrated assay, which included the newly integrated expression profiles. Based on the results of integration, we performed PCA dimensionality reduction using RunPCA on the variable features employed for integration. This was followed by UMAP dimensionality reduction using RunUMAP with parameters -dims 1:20, -n.neighbors 50. For unsupervised clustering, we executed the FindNeighbors function using default parameters, and FindClusters with -resolution 0.4.
Assembly: GRCm39
Supplementary files format and content: Tab-separated value files and matrix files
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| Submission date |
Apr 02, 2024 |
| Last update date |
May 01, 2024 |
| Contact name |
Xu Xiaopeng |
| E-mail(s) |
xu_xiaopeng@gzlab.ac.cn
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| Organization name |
Guangzhou National Laboratory
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| Street address |
96 Xingdao South Road, Guangzhou International Bio Island, Haizhu District
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| City |
Guangzhou |
| ZIP/Postal code |
510320 |
| Country |
China |
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| Platform ID |
GPL24247 |
| Series (1) |
| GSE263087 |
Regulatory Functional Landscape of the HMX1 Gene for Normal Ear Development [scRNA-seq] |
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| Relations |
| BioSample |
SAMN40731344 |
| SRA |
SRX24137164 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM8185650_GFP_E10.5_feature_bc_matrix.tar.gz |
50.6 Mb |
(ftp)(http) |
TAR |
SRA Run Selector |
| Raw data are available in SRA |
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