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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Aug 27, 2023 |
| Title |
SpiB replicate1 |
| Sample type |
SRA |
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| Source name |
Thymic hematopoietic cells
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| Organism |
Mus musculus |
| Characteristics |
cell type: Thymic hematopoietic cells
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| Extracted molecule |
total RNA |
| Extraction protocol |
Thymi from two 15wk old Spib+/– and two Spib–/– chimeras were surgically removed following perfusion, then trimmed of fat and connective tissues, and placed in PBS on ice. Single cell suspensions were prepared by mechanical dissociation of the thymi through a 40μm strainer using a syringe plunger. Cells were washed with 10 ml MACS buffer. 25 million cells per sample were first stained with 5 μl anti-mouse CD16/CD32 in 50 μl labeling buffer (PBS + 1% BSA) for 10 min at 4 °C, then an antibody mix containing both 5’ hashing antibodies and flow cytometry antibodies in labeling buffer was added to complete to ~100 μl total volume, and cells were incubated for 30 min at 4 °C. Cells were then washed 3 times with 5 ml labeling buffer. 10 minutes before each sample was sorted, it was incubated with a viability dye for 5 min at room temperature, then washed and resuspended in PBS + 2% BSA. CD45.1+CD3– (CD4/CD8DP)–MHCIImid-high cells were sorted into PBS supplemented with 50% FBS and then washed and resuspended in PBS + 0.04% BSA, counted, and volume was adjusted to a concentration of 1,000,000 cells/ml. Each two non-replicate samples were combined evenly into one pool.
The 10x Genomics platform loading and library preparation were done according to the manufacturer’s instructions, as previously described (10x Genomics) and were sequenced with paired-end 100-bp reads on an Illumina NovaSeq6000.
5' single cell RNA-seq
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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 |
Processing was done using the Cell Ranger ‘multi’ analysis pipeline (10x Genomics version 7.0), with the reference data file refdata-gex-mm10-2020-A.
Cell hashing oligo sequence for replicate1_KO: ACCCACCAGTAAGAC, for replicate1_HET:GGTCGAGAGCATTCA, for replicate2_KO:AAAGCATTCTTCACG, for replicate2_HET:CTTGCCGCATGTCAT
Assembly: mm10
Supplementary files format and content: Filtered feature-barcode matrix from cellranger
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| Submission date |
Jul 06, 2023 |
| Last update date |
Aug 27, 2023 |
| Contact name |
Jakub Abramson |
| E-mail(s) |
jakub.abramson@weizmann.ac.il
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| Organization name |
Weizmann Institute of Science
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| Department |
Department of Immunology
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| Street address |
234 Herzl st.
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| City |
Rehovot |
| ZIP/Postal code |
7610001 |
| Country |
Israel |
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| Platform ID |
GPL24247 |
| Series (2) |
| GSE236075 |
Thymic mimetic cells function beyond self-tolerance |
| GSE236669 |
Thymic mimetic cells function beyond self-tolerance [scRNA-Seq Spib chimeras] |
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| Relations |
| BioSample |
SAMN36345337 |
| SRA |
SRX20928465 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM7567343_barcodes1.tsv.gz |
65.8 Kb |
(ftp)(http) |
TSV |
| GSM7567343_features1.tsv.gz |
284.1 Kb |
(ftp)(http) |
TSV |
| GSM7567343_matrix1.mtx.gz |
77.2 Mb |
(ftp)(http) |
MTX |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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