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| Status |
Public on Apr 14, 2020 |
| Title |
Males |
| Sample type |
SRA |
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| Source name |
Pituitary
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| Organism |
Danio rerio |
| Characteristics |
genotype: Sox17:CreERT2;ubi:loxP-EGFP-loxP-mCherry
treatment: 4OH-Tamoxifen
tissue: Pituitary
age: 6 months
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| Growth protocol |
6months old animals (Males and Male1) and 3month old animal for transplant
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
Single cell was partitioned through Chromium platform (10X Genomic) which encapsulate each cell in individual GEMs. The cell lysis, polyA pull down, tagging with cell barcode, and RNA:DNA hybrid synthesis were performed in the GEMs. The version 2 of Chromium Single Cell 3' Reagent Kits was used for this RNA extraction step.
After cell-barcoding the extracted RNA, the GEMs were lysis for further fragmentation and ligation of illumina P5 and P7 ends with the version 2 of Chromium Single Cell 3' Reagent Kits. Two size selections were performed for long fragments (> 1000 bps) and short fragments (< 1000 bps) before fragmentation and after finalization respectively.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 4000 |
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| Description |
fresh 5uM 4-oh tamoxifen added at 4 hours post fertilization embryos, washed after 2h
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| Data processing |
10X Genomic Cellranger v3.0.0 was used for demultiplexing, alignment, and raw count matrix generation. The alignment was performed against in house built transcriptome index containing transgenic sequences (EGFP, mCherry, and TagBFP). QC step for the library was performed along with the Cellranger alignment process.
The raw count matrices were filtered for genes observed in at least 5 cells and cells that had at least 500 unique genes expressed.
To reduce the batch effects caused by the different strategies, we aligned the scaled data based on the CCA analysis by R package Seurat v2.4.0 to supress the variance introduced between libraries to generate an aggregated library.
The aggregated library was further filtered less than 0.04 mitochondrial gene expression and with 200 to 2500 unique genes.
With the aligned CCA space, we found cluster by Louvain clustering with 0.1 resolution and performed low dimensional embedding by UMAP through functions in Seurat. The assignment of cluster identity was done by examine the marker genes for different cell types reported in literatures.
Genome_build: GRCz11
Supplementary_files_format_and_content: tab-delimited file with cell barcode information, tab-delimited file with gene names, sparse matrix file with raw count associated with previous two files
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| Submission date |
Apr 13, 2020 |
| Last update date |
Apr 15, 2020 |
| Contact name |
Gage Crump |
| E-mail(s) |
gcrump@med.usc.edu
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| Phone |
323-442-2693
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| Organization name |
USC
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| Department |
Stem Cell
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| Lab |
Crump Lab
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| Street address |
1425 San Pablo St.
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| City |
Los Angeles |
| State/province |
CA |
| ZIP/Postal code |
90033 |
| Country |
USA |
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| Platform ID |
GPL21741 |
| Series (1) |
| GSE148591 |
Single cell transcriptomic analysis of endoderm derived cell lineages contribution in pituitary |
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| Relations |
| BioSample |
SAMN14592713 |
| SRA |
SRX8111241 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4474603_M_Pituitary_Sox17creERUbiGtR.mtx.gz |
28.5 Mb |
(ftp)(http) |
MTX |
| GSM4474603_M_Pituitary_Sox17creERUbiGtR_barcodes.tsv.gz |
38.5 Kb |
(ftp)(http) |
TSV |
| GSM4474603_M_Pituitary_Sox17creERUbiGtR_features.tsv.gz |
301.3 Kb |
(ftp)(http) |
TSV |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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