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| Status |
Public on Apr 29, 2024 |
| Title |
Neo, mRNA-derived cDNA, bio rep1 |
| Sample type |
SRA |
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| Source name |
ear
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| Organism |
Mus musculus |
| Characteristics |
strain background: C57BL/6
tissue: ear
cell type: endothelial
library type: mRNA
antibodies/tags: none
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
We first dissociated mouse ears and MACS enriched CD31+ cells from adult (2 months), adult wounded (day 9 post wounding), and neonatal (P8) samples. The endothelial enriched samples were stained with CD31-Alexa488, NearIR live/dead, and Antibody Derived Tag directed against Icam1, Vcam1, Cxcr4, Flk1 (VEGFR2), and Cd45. We then FACS sorted CD31+, Cdh5CreERT2-Rtom+, and NearIR- live endothelial cells and loaded them into a Chromium Controller (10X Genomics).
Library was prepared using Chromium Single Cell 3ʹ v3 protocol (10x Genomics). Cellular mRNA and antibody-derived oligos were reverse-transcribed and indexed with a shared cellular barcode. Indexed cDNAs were then pooled and amplified by PCR according to the Chromium Single Cell 3’ v3 protocol (10x Genomics) and with specific primers amplifying the antibody-derived tags. SPRI bead size selection was performed in order to separate both the mRNA-derived cDNA (>300bp) and the antibody-derived tagged cDNAs (180bp). For the mRNA derived cDNA library preparation, we further proceeded according to the manufacturer’s instructions for Single Cell 3’ v3 protocol (10x Genomics). For antibody-derived tagged library, we used the QuantaBio Hifi MasterMix following Biolegend's TotalSeqA protocol.
Following the final bead purification, all libraries were pooled as 95% mRNA library and 5% ADT library.
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| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina NextSeq 500 |
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| Description |
3' mRNA library: read1 file contains cell barcode and UMI; read2 file contains transcript
poly A RNA
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| Data processing |
library strategy: CITE-seq (Cellular Indexing of Transcriptomes and Epitopes by sequencing)
Pre-processing of sequencing results to generate count matrices (gene expression and ADT barcode counts) was performed using the 10x genomics Cell Ranger pipeline with default settings.The mm10 genome build was used and ADT sequences were concatenated and treated as Custom library.
Further processing was done with Seurat v3.0.2 (cell and gene filtering, clustering, differential gene expression analysis).
Assembly: mm10
Supplementary files format and content: 10x Genomics output files: barcodes.tsv.gz, features.tsv.gz, matrix.mtx.gz
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| Submission date |
Oct 26, 2022 |
| Last update date |
Apr 29, 2024 |
| Contact name |
Ziqing Liu |
| Organization name |
UNC-Chapel Hill
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| Department |
Biology
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| Lab |
Victoria Bautch
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| Street address |
416 Fordham Hall, University of North Carolina
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| City |
Chapel Hill |
| State/province |
NC |
| ZIP/Postal code |
27514 |
| Country |
USA |
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| Platform ID |
GPL19057 |
| Series (1) |
| GSE216594 |
Transcriptomic heterogeneity of endothelial cells during devleopmental angiogenesis |
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| Relations |
| BioSample |
SAMN31456412 |
| SRA |
SRX18029165 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM6681031_Neo1_barcodes.tsv.gz |
9.6 Kb |
(ftp)(http) |
TSV |
| GSM6681031_Neo1_features.tsv.gz |
272.8 Kb |
(ftp)(http) |
TSV |
| GSM6681031_Neo1_matrix.mtx.gz |
17.9 Mb |
(ftp)(http) |
MTX |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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